Enzymatically polymerized gelling dextrans

ABSTRACT

Compositions are disclosed herein comprising dextran that comprises (i) 87-93 wt % glucose linked at positions 1 and 6; (ii) 0.1-1.2 wt % glucose linked at positions 1 and 3; (iii) 0.1-0.7 wt % glucose linked at positions 1 and 4; (iv) 7.7-8.6 wt % glucose linked at positions 1, 3 and 6; and (v) about 0.4-1.7 wt % glucose linked at (a) positions 1, 2 and 6, or (b) positions 1, 4 and 6. Aqueous forms of this composition have enhanced viscosity profiles. Further disclosed are methods of using compositions comprising dextran, such as increasing the viscosity of an aqueous composition. Enzymatic reactions for producing dextran are also disclosed.

This application claims the benefit of U.S. Provisional Application No.62/075,460 (filed Nov. 5, 2014), which is incorporated herein byreference in its entirety.

FIELD OF INVENTION

The present disclosure is in the field of polysaccharides. For example,the disclosure pertains to certain dextran polymers, reactionscomprising glucosyltransferase enzymes that synthesize these polymers,and use of the polymers in various applications.

REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY

The official copy of the sequence listing is submitted electronicallyvia EFS-Web as an ASCII formatted sequence listing with a file named20151105_CL6294USNP_SequenceListing.txt created on Nov. 5, 2015, andhaving a size of 164 kilobytes and is filed concurrently with thespecification. The sequence listing contained in this ASCII-formatteddocument is part of the specification and is herein incorporated byreference in its entirety.

BACKGROUND

Driven by a desire to find new structural polysaccharides usingenzymatic syntheses or genetic engineering of microorganisms,researchers have discovered polysaccharides that are biodegradable andcan be made economically from renewably sourced feedstocks. One suchfamily of polysaccharides are alpha-glucans, which are polymerscomprising glucose monomers linked by alpha-glycosidic bonds.

Dextrans represent a family of complex, branched alpha-glucans generallycomprising chains of alpha-1,6-linked glucose monomers, with periodicside chains (branches) linked to the straight chains byalpha-1,3-linkage (loan et al., Macromolecules 33:5730-5739). Productionof dextrans is typically done through fermentation of sucrose withbacteria (e.g., Leuconostoc or Streptococcus species), where sucroseserves as the source of glucose for dextran polymerization (Naessens etal., J. Chem. Technol. Biotechnol. 80:845-860; Sarwat et al., Int. J.Biol. Sci. 4:379-386; Onilude et al., Int. Food Res. J. 20:1645-1651).Although dextrans are used in several applications given their highsolubility in water (e.g., adjuvants, stabilizers), this high solubilitycan negatively affect their general utility as thickening agents inhydrocolloid applications.

Thus, there is interest in developing new, higher viscosity dextranpolymers that are more amenable to high viscosity applications. In turn,there is also interest in identifying glucosyltransferase enzymes thatcan synthesize such dextran polymers.

SUMMARY OF INVENTION

In one embodiment, the disclosure concerns a composition comprisingdextran that comprises:

(i) about 87-93 wt % glucose linked at positions 1 and 6;

(ii) about 0.1-1.2 wt % glucose linked at positions 1 and 3;

(iii) about 0.1-0.7 wt % glucose linked at positions 1 and 4;

(iv) about 7.7-8.6 wt % glucose linked at positions 1, 3 and 6; and

(v) about 0.4-1.7 wt % glucose linked at:

-   -   (a) positions 1, 2 and 6, or    -   (b) positions 1, 4 and 6;        wherein the weight-average molecular weight (Mw) of the dextran        is about 50-200 million Daltons, the z-average radius of        gyration of the dextran is about 200-280 nm, and the dextran        optionally is not a product of a Leuconostoc mesenteroides        glucosyltransferase enzyme.

In another embodiment, the dextran comprises: (i) about 89.5-90.5 wt %glucose linked at positions 1 and 6; (ii) about 0.4-0.9 wt % glucoselinked at positions 1 and 3; (iii) about 0.3-0.5 wt % glucose linked atpositions 1 and 4; (iv) about 8.0-8.3 wt % glucose linked at positions1, 3 and 6; and (v) about 0.7-1.4 wt % glucose linked at: (a) positions1, 2 and 6, or (b) positions 1, 4 and 6.

In another embodiment, the dextran comprises chains (long chains) linkedtogether within a branching structure, wherein said chains are similarin length and comprise substantially alpha-1,6-glucosidic linkages. Theaverage length of the chains is about 10-50 monomeric units in anotherembodiment.

In another embodiment, the dextran is a product of a glucosyltransferaseenzyme comprising an amino acid sequence that is at least 90% identicalto SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:9, SEQ ID NO:13, orSEQ ID NO:17.

In another embodiment, the composition is an aqueous composition havinga viscosity of at least about 25 cPs.

In another embodiment, the Mw of the dextran is about 80-120 millionDaltons.

In another embodiment, the z-average radius of gyration of the dextranis about 230-250 nm.

In another embodiment, the composition is in the form of a food product,personal care product, pharmaceutical product, household product, orindustrial product. In another embodiment, the composition is in theform of a confectionery.

In another embodiment, the disclosure concerns a method for increasingthe viscosity of an aqueous composition. This method comprisescontacting at least one dextran compound as disclosed herein with anaqueous composition. The contacting step in this method results inincreasing the viscosity of the aqueous composition, in comparison tothe viscosity of the aqueous composition before the contacting step.

In another embodiment, the disclosure concerns a method of treating amaterial. This method comprises contacting a material with an aqueouscomposition comprising at least one dextran compound disclosed herein.

In another embodiment, the disclosure concerns an enzymatic reactioncomprising water, sucrose and a glucosyltransferase enzyme comprising anamino acid sequence that is at least 90% identical to SEQ ID NO:1, SEQID NO:2, SEQ ID NO:5, SEQ ID NO:9, SEQ ID NO:13, or SEQ ID NO:17,wherein the glucosyltransferase enzyme synthesizes a dextran compound asdisclosed herein.

In another embodiment, the disclosure concerns a method of producingdextran comprising the step of contacting at least water, sucrose, and aglucosyltransferase enzyme comprising an amino acid sequence that is atleast 90% identical to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:5, SEQ IDNO:9, SEQ ID NO:13, or SEQ ID NO:17, thereby producing dextran asdisclosed herein. This dextran can optionally be isolated.

In another embodiment, the viscosity of the dextran produced in themethod is increased by decreasing the amount of sucrose in step (a).

BRIEF DESCRIPTION OF THE DRAWINGS AND SEQUENCES

FIG. 1: HPLC analysis of sucrose consumption by a glucosyltransferasereaction comprising 100 g/L sucrose and a 0768 gtf (SEQ ID NO:1). Referto Example 2.

FIG. 2A: Map of plasmid pZZHB583 used to express 2919 gtf (SEQ ID NO:5)in B. subtilis. Refer to Example 3.

FIG. 2B: Map of plasmid pZZHB582 used to express 2918 gtf (SEQ ID NO:9)in B. subtilis. Refer to Example 4.

FIG. 2C: Map of plasmid pZZHB584 used to express 2920 gtf (SEQ ID NO:13)in B. subtilis. Refer to Example 5.

FIG. 2D: Map of plasmid pZZHB585 used to express 2921 (SEQ ID NO:17) gtfin B. subtilis. Refer to Example 6.

FIG. 3: HPLC analysis of sucrose consumption by a reaction comprising acommercially available dextran sucrase. Refer to Example 7.

TABLE 1 Summary of Nucleic Acid and Protein SEQ ID Numbers Nucleic acidProtein SEQ ID SEQ ID Description NO. NO. “0768 gtf”, Leuconostocpseudomesenteroides. 1 Mature form of GENBANK Identification No. (1447aa) 497964659. “0768 gtf”, Leuconostoc pseudomesenteroides. 2 Matureform of GENBANK Identification No. (1457 aa) 497964659, but including astart methionine and additional N- and C-terminal amino acids. WciGtf1,Weissella cibaria. Full length form 3 4 comprising signal sequence.GENBANK Accession (4347 (1448 aa) No. ZP_08417432 (amino acid sequence).bases) “2919 gtf”, Weissella cibaria. Mature form 5 of GENBANKIdentification No. ZP_08417432. (1422 aa) “2919 gtf”, Weissella cibaria.Sequence 6 optimized for expression in B. subtilis. (4269 Encodes 2919gtf with a heterologous signal bases) sequence and additional N-terminalamino acids. LfeGtf1, Lactobacillus fermentum. Full length 7 8 formcomprising signal sequence. GENBANK (4392 (1463 aa) Accession No.AAU08008 (amino acid sequence). bases) “2918 gtf”, Lactobacillusfermentum. Mature 9 form of GENBANK Identification No. AAU08008. (1426aa) “2918 gtf”, Lactobacillus fermentum. Sequence 10 optimized forexpression in B. subtilis. (4281 Encodes 2918 gtf with a heterologoussignal bases) sequence and additional N-terminal amino acids. SsoGtf4,Streptococcus sobrinus. Full length 11 12 form comprising signalsequence. GENBANK (4521 (1506 aa) Accession No. AAX76986 (amino acidsequence). bases) “2920 gtf”, Streptococcus sobrinus. Mature 13 form ofGENBANK Identification No. AAX76986. (1465 aa) “2920 gtf”, Streptococcussobrinus. Sequence 14 optimized for expression in B. subtilis. (4398Encodes 2920 gtf with a heterologous signal bases) sequence andadditional N-terminal amino acids. SdoGtf7, Streptococcus downei. Fulllength form 15 16 comprising signal sequence. GENBANK Accession (4360(1453 aa) No. ZP_08549987.1 (amino acid sequence). bases) “2921 gtf”,Streptococcus downei. Mature form 17 of GENBANK Identification No.ZP_08549987.1. (1409 aa) “2921 gtf”, Streptococcus downei. Sequence 18optimized for expression in B. subtilis. (4230 Encodes 2921 gtf with aheterologous signal bases) sequence and additional N-terminal aminoacids.

DETAILED DESCRIPTION

The disclosures of all cited patent and non-patent literature areincorporated herein by reference in their entirety.

Unless otherwise disclosed, the terms “a” and “an” as used herein areintended to encompass one or more (i.e., at least one) of a referencedfeature.

The term “glucan” herein refers to a polysaccharide of D-glucosemonomers that are linked by glucosidic linkages, which are a type ofglycosidic linkage. An “alpha-glucan” herein refers to a glucan in whichthe constituent D-glucose monomers are alpha-D-glucose monomers.

The terms “dextran”, “dextran polymer”, “dextran compound” and the likeare used interchangeably herein and refer to complex, branchedalpha-glucans generally comprising chains of substantially (mostly)alpha-1,6-linked glucose monomers, with side chains (branches) linkedmainly by alpha-1,3-linkage. The term “gelling dextran” herein refers tothe ability of one or more dextrans disclosed herein to form a viscoussolution or gel-like composition (i) during enzymatic dextran synthesisand, optionally, (ii) when such synthesized dextran is isolated(e.g., >90% pure) and then placed in an aqueous composition.

Dextran “long chains” herein can comprise “substantially [or mostly]alpha-1,6-glucosidic linkages”, meaning that they can have at leastabout 98.0% alpha-1,6-glucosidic linkages in some aspects. Dextranherein can comprise a “branching structure” (branched structure) in someaspects. It is contemplated that in this structure, long chains branchfrom other long chains, likely in an iterative manner (e.g., a longchain can be a branch from another long chain, which in turn can itselfbe a branch from another long chain, and so on). It is contemplated thatlong chains in this structure can be “similar in length”, meaning thatthe length (DP [degree of polymerization]) of at least 70% of all thelong chains in a branching structure is within plus/minus 30% of themean length of all the long chains of the branching structure.

Dextran in some embodiments can also comprise “short chains” branchingfrom the long chains, typically being one to three glucose monomers inlength, and comprising less than about 10% of all the glucose monomersof a dextran polymer. Such short chains typically comprise alpha-1,2-,alpha-1,3-, and/or alpha-1,4-glucosidic linkages (it is believed thatthere can also be a small percentage of such non-alpha-1,6 linkages inlong chains in some aspects).

The terms “glycosidic linkage” and “glycosidic bond” are usedinterchangeably herein and refer to the covalent bond that joins acarbohydrate molecule to another carbohydrate molecule. The terms“glucosidic linkage” and “glucosidic bond” are used interchangeablyherein and refer to a glycosidic linkage between two glucose molecules.The term “alpha-1,6-glucosidic linkage” as used herein refers to thecovalent bond that joins alpha-D-glucose molecules to each other throughcarbons 1 and 6 on adjacent alpha-D-glucose rings. The term“alpha-1,3-glucosidic linkage” as used herein refers to the covalentbond that joins alpha-D-glucose molecules to each other through carbons1 and 3 on adjacent alpha-D-glucose rings. The term“alpha-1,2-glucosidic linkage” as used herein refers to the covalentbond that joins alpha-D-glucose molecules to each other through carbons1 and 2 on adjacent alpha-D-glucose rings. The term“alpha-1,4-glucosidic linkage” as used herein refers to the covalentbond that joins alpha-D-glucose molecules to each other through carbons1 and 4 on adjacent alpha-D-glucose rings. Herein, “alpha-D-glucose”will be referred to as “glucose.” All glucosidic linkages disclosedherein are alpha-glucosidic linkages, except where otherwise noted.

“Glucose (glucose monomers) linked at positions 1 and 6” herein refersto a glucose monomer of dextran in which only carbons 1 and 6 of theglucose monomer are involved in respective glucosidic linkages with twoadjacent glucose monomers. This definition likewise applies to glucose(i) “linked at positions 1 and 3”, and (ii) “linked at positions 1 and4”, taking into account, accordingly, the different carbon positionsinvolved in each respective linkage.

“Glucose (glucose monomers) linked at positions 1, 3 and 6” hereinrefers to a glucose monomer of dextran in which carbons 1, 3 and 6 ofthe glucose monomer are involved in respective glucosidic linkages withthree adjacent glucose monomers. A glucose linked only at positions 1, 3and 6 is a branch point. This definition likewise applies to glucoselinked at (i) positions 1, 2 and 6, and (ii) positions 1, 4 and 6, buttaking into account, accordingly, the different carbon positionsinvolved in each respective linkage.

Glucose positions (glucose carbon positions) 1, 2, 3, 4 and 6 herein areas known in the art (depicted in the following structure):

The glycosidic linkage profile of a dextran herein can be determinedusing any method known in the art. For example, a linkage profile can bedetermined using methods that use nuclear magnetic resonance (NMR)spectroscopy (e.g., ¹³C NMR or ¹H NMR). These and other methods that canbe used are disclosed in Food Carbohydrates: Chemistry, PhysicalProperties, and Applications (S. W. Cui, Ed., Chapter 3, S. W. Cui,Structural Analysis of Polysaccharides, Taylor & Francis Group LLC, BocaRaton, Fla., 2005), which is incorporated herein by reference.

The term “sucrose” herein refers to a non-reducing disaccharide composedof an alpha-D-glucose molecule and a beta-D-fructose molecule linked byan alpha-1,2-glycosidic bond. Sucrose is known commonly as table sugar.

The “molecular weight” of dextran herein can be represented asnumber-average molecular weight (Mn) or as weight-average molecularweight (Mw), the units of which are in Daltons or grams/mole.Alternatively, molecular weight can be represented as DPw (weightaverage degree of polymerization) or DPn (number average degree ofpolymerization). Various means are known in the art for calculatingthese molecular weight measurements such as with high-pressure liquidchromatography (HPLC), size exclusion chromatography (SEC), or gelpermeation chromatography (GPC).

The term “radius of gyration” (Rg) herein refers to the mean radius ofdextran, and is calculated as the root-mean-square distance of a dextranmolecule's components (atoms) from the molecule's center of gravity. Rgcan be provided in Angstrom or nanometer (nm) units, for example. The“z-average radius of gyration” of dextran herein refers to the Rg ofdextran as measured using light scattering (e.g., MALS). Methods formeasuring z-average Rg are known and can be used herein, accordingly.For example, z-average Rg can be measured as disclosed in U.S. Pat. No.7,531,073, U.S. Patent Appl. Publ. Nos. 2010/0003515 and 2009/0046274,Wyatt (Anal. Chim. Acta 272:1-40), and Mori and Barth (Size ExclusionChromatography, Springer-Verlag, Berlin, 1999), all of which areincorporated herein by reference.

The terms “glucosyltransferase enzyme”, “gtf enzyme”, “gtf enzymecatalyst”, “gtf”, “glucansucrase” and the like are used interchangeablyherein. The activity of a gtf enzyme herein catalyzes the reaction ofthe substrate sucrose to make the products glucan and fructose. A gtfenzyme that produces a dextran (a type of glucan) can also be referredto as a dextransucrase. Other products (byproducts) of a gtf reactioncan include glucose (where glucose is hydrolyzed from the glucosyl-gtfenzyme intermediate complex), and various soluble oligosaccharides(e.g., DP2-DP7) such as leucrose, Wild type forms of glucosyltransferaseenzymes generally contain (in the N-terminal to C-terminal direction) asignal peptide, a variable domain, a catalytic domain, and aglucan-binding domain. A gtf herein is classified under the glycosidehydrolase family 70 (GH70) according to the CAZy (Carbohydrate-ActiveEnZymes) database (Cantarel et al., Nucleic Acids Res. 37:D233-238,2009).

The terms “glucosyltransferase catalytic domain” and “catalytic domain”are used interchangeably herein and refer to the domain of aglucosyltransferase enzyme that provides glucan-producing activity tothe glucosyltransferase enzyme.

The terms “gtf reaction”, “gtf reaction solution”, “glucosyltransferasereaction”, “enzymatic reaction”, “dextran synthesis reaction”, “dextranreaction” and the like are used interchangeably herein and refer to areaction that is performed by a glucosyltransferase enzyme. A gtfreaction as used herein generally refers to a reaction initiallycomprising at least one active glucosyltransferase enzyme in a solutioncomprising sucrose and water, and optionally other components. Othercomponents that can be in a gtf reaction after it has commenced includefructose, glucose, soluble oligosaccharides (e.g., DP2-DP7) such asleucrose, and dextran products. It is in a gtf reaction where the stepof contacting water, sucrose and a glucosyltransferase enzyme isperformed. The term “under suitable gtf reaction conditions” as usedherein, refers to gtf reaction conditions that support conversion ofsucrose to dextran via glucosyltransferase enzyme activity. A gtfreaction herein is not naturally occurring.

A “control” gtf reaction as used herein can refer to a reaction using aglucosyltransferase not comprising an amino acid sequence that is atleast 90% identical to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:5, SEQ IDNO:9, SEQ ID NO:13, or SEQ ID NO:17. All the other features (e.g.,sucrose concentration, temperature, pH, time) of a control reactionsolution can be the same as the reaction to which it is being compared.

The “percent dry solids” of a gtf reaction refers to the wt % of all thesugars in a gtf reaction. The percent dry solids of a gtf reaction canbe calculated, for example, based on the amount of sucrose used toprepare the reaction.

The “yield” of dextran by a gtf reaction herein represents the weight ofdextran product expressed as a percentage of the weight of sucrosesubstrate that is converted in the reaction. For example, if 100 g ofsucrose in a reaction solution is converted to products, and 10 g of theproducts is dextran, the yield of the dextran would be 10%. This yieldcalculation can be considered as a measure of selectivity of thereaction toward dextran.

The terms “percent by volume”, “volume percent”, “vol %”, “v/v %” andthe like are used interchangeably herein. The percent by volume of asolute in a solution can be determined using the formula: [(volume ofsolute)/(volume of solution)]×100%.

The terms “percent by weight”, “weight percentage (wt %)”,“weight-weight percentage (% w/w)” and the like are used interchangeablyherein. Percent by weight refers to the percentage of a material on amass basis as it is comprised in a composition, mixture, or solution.

The term “increased” as used herein can refer to a quantity or activitythat is at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%,12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 50%, 100%, or 200% morethan the quantity or activity for which the increased quantity oractivity is being compared. The terms “increased”, “elevated”,“enhanced”, “greater than”, “improved” and the like are usedinterchangeably herein.

The terms “polynucleotide”, “polynucleotide sequence”, and “nucleic acidsequence” are used interchangeably herein. These terms encompassnucleotide sequences and the like. A polynucleotide may be a polymer ofDNA or RNA that is single- or double-stranded, that optionally containssynthetic, non-natural or altered nucleotide bases. A polynucleotide maybe comprised of one or more segments of cDNA, genomic DNA, syntheticDNA, or mixtures thereof.

The term “gene” as used herein refers to a DNA polynucleotide sequencethat expresses an RNA (RNA is transcribed from the DNA polynucleotidesequence) from a coding region (coding sequence), which RNA can be amessenger RNA (encoding a protein) or a non-protein-coding RNA. A genemay refer to the coding region alone, or may include regulatorysequences upstream and/or downstream to the coding region (e.g.,promoters, 5′-untranslated regions, 3′-transcription terminatorregions). A coding region encoding a protein can alternatively bereferred to herein as an “open reading frame” (ORF). A gene that is“native” or “endogenous” refers to a gene as found in nature with itsown regulatory sequences; such a gene is located in its natural locationin the genome of a host cell. A “chimeric” gene refers to any gene thatis not a native gene, comprising regulatory and coding sequences thatare not found together in nature (i.e., the regulatory and codingregions are heterologous with each other). Accordingly, a chimeric genemay comprise regulatory sequences and coding sequences that are derivedfrom different sources, or regulatory sequences and coding sequencesderived from the same source, but arranged in a manner different thanthat found in nature. A “foreign” or “heterologous” gene refers to agene that is introduced into a host organism by gene transfer. Foreigngenes can comprise native genes inserted into a non-native organism,native genes introduced into a new location within the native host, orchimeric genes. Polynucleotide sequences in certain embodimentsdisclosed herein are heterologous. A “transgene” is a gene that has beenintroduced into the genome by a transformation procedure. A“codon-optimized” open reading frame has its frequency of codon usagedesigned to mimic the frequency of preferred codon usage of the hostcell.

The term “recombinant” or “heterologous” as used herein refers to anartificial combination of two otherwise separated segments of sequence,e.g., by chemical synthesis or by the manipulation of isolated segmentsof nucleic acids by genetic engineering techniques. The terms“recombinant”, “transgenic”, “transformed”, “engineered” or “modifiedfor exogenous gene expression” are used interchangeably herein.

A native amino acid sequence or polynucleotide sequence is naturallyoccurring, whereas a non-native amino acid sequence or polynucleotidesequence does not occur in nature.

“Regulatory sequences” as used herein refer to nucleotide sequenceslocated upstream of a gene's transcription start site (e.g., promoter),5′ untranslated regions, and 3′ non-coding regions, and which mayinfluence the transcription, processing or stability, or translation ofan RNA transcribed from the gene. Regulatory sequences herein mayinclude promoters, enhancers, silencers, 5′ untranslated leadersequences, introns, polyadenylation recognition sequences, RNAprocessing sites, effector binding sites, stem-loop structures, andother elements involved in regulation of gene expression. One or moreregulatory elements herein may be heterologous to a coding regionherein.

Methods for preparing recombinant constructs/vectors herein can followstandard recombinant DNA and molecular cloning techniques as describedby J. Sambrook and D. Russell (Molecular Cloning: A Laboratory Manual,3rd Edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor,N.Y., 2001); T. J. Silhavy et al. (Experiments with Gene Fusions, ColdSpring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1984); and F.M. Ausubel et al. (Short Protocols in Molecular Biology, 5th Ed. CurrentProtocols, John Wiley and Sons, Inc., NY, 2002).

The terms “sequence identity” or “identity” as used herein with respectto polynucleotide or polypeptide sequences refer to the nucleic acidbases or amino acid residues in two sequences that are the same whenaligned for maximum correspondence over a specified comparison window.Thus, “percentage of sequence identity” or “percent identity” refers tothe value determined by comparing two optimally aligned sequences over acomparison window, wherein the portion of the polynucleotide orpolypeptide sequence in the comparison window may comprise additions ordeletions (i.e., gaps) as compared to the reference sequence (which doesnot comprise additions or deletions) for optimal alignment of the twosequences. The percentage is calculated by determining the number ofpositions at which the identical nucleic acid base or amino acid residueoccurs in both sequences to yield the number of matched positions,dividing the number of matched positions by the total number ofpositions in the window of comparison and multiplying the results by 100to yield the percentage of sequence identity. It would be understoodthat, when calculating sequence identity between a DNA sequence and anRNA sequence, T residues of the DNA sequence align with, and can beconsidered “identical” with, U residues of the RNA sequence. Forpurposes of determining percent complementarity of first and secondpolynucleotides, one can obtain this by determining (i) the percentidentity between the first polynucleotide and the complement sequence ofthe second polynucleotide (or vice versa), for example, and/or (ii) thepercentage of bases between the first and second polynucleotides thatwould create canonical Watson and Crick base pairs.

The Basic Local Alignment Search Tool (BLAST) algorithm, which isavailable online at the National Center for Biotechnology Information(NCBI) website, may be used, for example, to measure percent identitybetween or among two or more of the polynucleotide sequences (BLASTNalgorithm) or polypeptide sequences (BLASTP algorithm) disclosed herein.Alternatively, percent identity between sequences may be performed usinga Clustal algorithm (e.g., ClustalW, ClustalV, or Clustal-Omega). Formultiple alignments using a Clustal method of alignment, the defaultvalues may correspond to GAP PENALTY=10 and GAP LENGTH PENALTY=10.Default parameters for pairwise alignments and calculation of percentidentity of protein sequences using a Clustal method may be KTUPLE=1,GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. For nucleic acids, theseparameters may be KTUPLE=2, GAP PENALTY=5, WINDOW=4 and DIAGONALSSAVED=4. Alternatively still, percent identity between sequences may beperformed using an EMBOSS algorithm (e.g., needle) with parameters suchas GAP OPEN=10, GAP EXTEND=0.5, END GAP PENALTY=false, END GAP OPEN=10,END GAP EXTEND=0.5 using a BLOSUM matrix (e.g., BLOSUM62).

Various polypeptide amino acid sequences and polynucleotide sequencesare disclosed herein as features of certain embodiments. Variants ofthese sequences that are at least about 70-85%, 85-90%, or 90%-95%identical to the sequences disclosed herein can be used. Alternatively,a variant amino acid sequence or polynucleotide sequence can have atleast 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%,83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98% or 99% identity with a sequence disclosed herein. The variantamino acid sequence or polynucleotide sequence may have the samefunction/activity of the disclosed sequence, or at least about 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% ofthe function/activity of the disclosed sequence. Any polypeptide aminoacid sequence disclosed herein not beginning with a methionine cantypically further comprise at least a start-methionine at the N-terminusof the amino acid sequence. Any polypeptide amino acid sequencedisclosed herein beginning with a methionine can optionally beconsidered without this methionine residue (i.e., a polypeptide sequencecan be referred to in reference to the position-2 residue to theC-terminal residue of the sequence).

The term “isolated” as used herein refers to any cellular component thathas been completely or partially purified from its native source (e.g.,an isolated polynucleotide or polypeptide molecule). In some instances,an isolated polynucleotide or polypeptide molecule is part of a greatercomposition, buffer system or reagent mix. For example, an isolatedpolynucleotide or polypeptide molecule can be comprised within a cell ororganism in a heterologous manner. Another example is an isolatedglucosyltransferase enzyme or reaction. “Isolated” herein can alsocharacterize a dextran compound. As such, dextran compounds of thepresent disclosure are synthetic, man-made compounds, and/or exhibitproperties not believed to naturally occur.

An “aqueous composition” herein has a liquid component that comprises atleast about 10 wt % water, for example. Examples of aqueous compositionsinclude mixtures, solutions, dispersions (e.g., colloidal dispersions),suspensions and emulsions, for example. Aqueous compositions in certainembodiments comprise dextran that is dissolved in the aqueouscomposition (i.e., in solution, and typically has viscosity).

As used herein, the term “colloidal dispersion” refers to aheterogeneous system having a dispersed phase and a dispersion medium,i.e., microscopically dispersed insoluble particles are suspendedthroughout another substance (e.g., an aqueous composition such as wateror aqueous solution). An example of a colloidal dispersion herein is ahydrocolloid. All, or a portion of, the particles of a colloidaldispersion such as a hydrocolloid can comprise certain dextran compoundsof the present disclosure. The terms “dispersant” and “dispersion agent”are used interchangeably herein to refer to a material that promotes theformation and/or stabilization of a dispersion.

The terms “hydrocolloid” and “hydrogel” are used interchangeably herein.A hydrocolloid refers to a colloid system in which water is thedispersion medium.

The term “aqueous solution” herein refers to a solution in which thesolvent comprises water. An aqueous solution can serve as a dispersantin certain aspects herein. Dextran compounds in certain embodiments canbe dissolved, dispersed, or mixed within an aqueous solution.

The terms “dispersant”, “dispersion agent” and the like are usedinterchangeably herein to refer to a material that promotes theformation and stabilization of a dispersion of one substance in another.A “dispersion” herein refers to an aqueous composition comprising one ormore particles (e.g., any ingredient of a personal care product,pharmaceutical product, food product, household product, or industrialproduct disclosed herein) that are scattered, or uniformly scattered,throughout the aqueous composition.

The term “viscosity” as used herein refers to the measure of the extentto which a fluid or an aqueous composition such as a hydrocolloidresists a force tending to cause it to flow. Various units of viscositythat can be used herein include centipoise (cPs) and Pascal-second(Pa·s). A centipoise is one one-hundredth of a poise; one poise is equalto 0.100 kg·m⁻¹·s⁻¹. Thus, the terms “viscosity modifier”,“viscosity-modifying agent” and the like as used herein refer toanything that can alter/modify the viscosity of a fluid or aqueouscomposition.

The term “shear thinning behavior” as used herein refers to a decreasein the viscosity of an aqueous composition as shear rate increases. Theterm “shear thickening behavior” as used herein refers to an increase inthe viscosity of an aqueous composition as shear rate increases. “Shearrate” herein refers to the rate at which a progressive shearingdeformation is applied to an aqueous composition. A shearing deformationcan be applied rotationally.

The term “contacting” as used herein with respect to methods ofincreasing the viscosity of an aqueous composition refers to any actionthat results in bringing together an aqueous composition with a dextran.Contacting can be performed by any means known in the art, such asdissolving, mixing, shaking, or homogenization, for example.

The terms “confectionery”, “confection”, “sweets”, “sweetmeat”, “candy”and the like are used interchangeably herein. A confectionary refers toany flavored food product having a sweet taste, the consistency of whichmay be hard or soft, which is typically consumed by sucking and/or bychewing within the oral cavity. A confectionary can contain sugar orotherwise be sugar-free.

The terms “fabric”, “textile”, “cloth” and the like are usedinterchangeably herein to refer to a woven material having a network ofnatural and/or artificial fibers. Such fibers can be thread or yarn, forexample.

A “fabric care composition” herein is any composition suitable fortreating fabric in some manner. Examples of such a composition includelaundry detergents and fabric softeners.

The terms “heavy duty detergent”, “all-purpose detergent” and the likeare used interchangeably herein to refer to a detergent useful forregular washing of white and colored textiles at any temperature. Theterms “low duty detergent” or “fine fabric detergent” are usedinterchangeably herein to refer to a detergent useful for the care ofdelicate fabrics such as viscose, wool, silk, microfiber or other fabricrequiring special care. “Special care” can include conditions of usingexcess water, low agitation, and/or no bleach, for example.

A “detergent composition” herein typically comprises at least onesurfactant (detergent compound) and/or at least one builder. A“surfactant” herein refers to a substance that tends to reduce thesurface tension of a liquid in which the substance is dissolved. Asurfactant may act as a detergent, wetting agent, emulsifier, foamingagent, and/or dispersant, for example.

The terms “anti-redeposition agent”, “anti-soil redeposition agent”,“anti-greying agent” and the like herein refer to agents that help keepsoils from redepositing onto clothing in laundry wash water after thesesoils have been removed, therefore preventing greying/discoloration oflaundry. Anti-redeposition agents can function by helping keep soildispersed in wash water and/or by blocking attachment of soil ontofabric surfaces.

An “oral care composition” herein is any composition suitable fortreating an soft or hard surface in the oral cavity such as dental(teeth) and/or gum surfaces.

The term “adsorption” herein refers to the adhesion of a compound (e.g.,dextran herein) to the surface of a material.

The terms “cellulase”, “cellulase enzyme” and the like are usedinterchangeably herein to refer to an enzyme that hydrolyzesbeta-1,4-D-glucosidic linkages in cellulose, thereby partially orcompletely degrading cellulose. Cellulase can alternatively be referredto as “beta-1,4-glucanase”, for example, and can have endocellulaseactivity (EC 3.2.1.4), exocellulase activity (EC 3.2.1.91), orcellobiase activity (EC 3.2.1.21). “Cellulose” refers to an insolublepolysaccharide having a linear chain of beta-1,4-linked D-glucosemonomeric units.

There is interest in developing new, high viscosity dextran polymers,which are more amenable to gelling applications. In turn, there is alsointerest in identifying glucosyltransferase enzymes that can synthesizesuch dextran polymers.

Embodiments of the present disclosure concern a composition comprising adextran that comprises:

-   -   (i) about 87-93 wt % glucose linked at positions 1 and 6;    -   (ii) about 0.1-1.2 wt % glucose linked at positions 1 and 3;    -   (iii) about 0.1-0.7 wt % glucose linked at positions 1 and 4;    -   (iv) about 7.7-8.6 wt % glucose linked at positions 1, 3 and 6;        and    -   (v) about 0.4-1.7 wt % glucose linked at: (a) positions 1, 2 and        6, or (b) positions 1, 4 and 6.        The weight-average molecular weight (Mw) and z-average radius of        gyration of such dextran is about 50-200 million Daltons and        about 200-280 nm, respectively. Also, such dextran optionally is        not a product of a Leuconostoc mesenteroides glucosyltransferase        enzyme.

An example of this composition is a glucosyltransferase reaction inwhich a dextran with the above linkage, weight and size profile issynthesized. Significantly, this dextran exhibits high viscosity inaqueous compositions, even at relatively low concentrations of thedextran. It is believed that this high viscosity profile is unique incomparison to viscosity profiles of previously disclosed dextranpolymers.

A dextran herein can comprise (i) about 87-93 wt % glucose linked onlyat positions 1 and 6; (ii) about 0.1-1.2 wt % glucose linked only atpositions 1 and 3; (iii) about 0.1-0.7 wt % glucose linked only atpositions 1 and 4; (iv) about 7.7-8.6 wt % glucose linked only atpositions 1, 3 and 6; and (v) about 0.4-1.7 wt % glucose linked only at:(a) positions 1, 2 and 6, or (b) positions 1, 4 and 6. In certainembodiments, a dextran can comprise (i) about 89.5-90.5 wt % glucoselinked only at positions 1 and 6; (ii) about 0.4-0.9 wt % glucose linkedonly at positions 1 and 3; (iii) about 0.3-0.5 wt % glucose linked onlyat positions 1 and 4; (iv) about 8.0-8.3 wt % glucose linked only atpositions 1, 3 and 6; and (v) about 0.7-1.4 wt % glucose linked only at:(a) positions 1, 2 and 6, or (b) positions 1, 4 and 6.

A dextran in some aspects of the present disclosure can comprise about87, 87.5, 88, 88.5, 89, 89.5, 90, 90.5, 91, 91.5, 92, 92.5, or 93 wt %glucose linked only at positions 1 and 6. There can be about 87-92.5,87-92, 87-91.5, 87-91, 87-90.5, 87-90, 87.5-92.5, 87.5-92, 87.5-91.5,87.5-91, 87.5-90.5, 87.5-90, 88-92.5, 88-92, 88-91.5, 88-91, 88-90.5,88-90, 88.5-92.5, 88.5-92, 88.5-91.5, 88.5-91, 88.5-90.5, 88.5-90,89-92.5, 89-92, 89-91.5, 89-91, 89-90.5, 89-90, 89.5-92.5, 89.5-92,89.5-91.5, 89.5-91, or 89.5-90.5 wt % glucose linked only at positions 1and 6, in some instances.

A dextran in some aspects can comprise about 0.1, 0.2, 0.3, 0.4, 0.5,0.6, 0.7, 0.8, 0.9, 1.0, 1.1, or 1.2 wt % glucose linked only atpositions 1 and 3. There can be about 0.1-1.2, 0.1-1.0, 0.1-0.8,0.3-1.2, 0.3-1.0, 0.3-0.8, 0.4-1.2, 0.4-1.0, 0.4-0.8, 0.5-1.2, 0.5-1.0,0.5-0.8, 0.6-1.2, 0.6-1.0, or 0.6-0.8 wt % glucose linked only atpositions 1 and 3, in some instances.

A dextran in some aspects can comprise about 0.1, 0.2, 0.3, 0.4, 0.5,0.6, or 0.7 wt % glucose linked only at positions 1 and 4. There can beabout 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1-0.4, 0.2-0.7, 0.2-0.6, 0.2-0.5,0.2-0.4, 0.3-0.7, 0.3-0.6, 0.3-0.5, or 0.3-0.4 wt % glucose linked onlyat positions 1 and 4, in some instances.

A dextran in some aspects can comprise about 7.7, 7.8, 7.9, 8.0, 8.1,8.2, 8.3, 8.4, 8.5, or 8.6 wt % glucose linked only at positions 1, 3and 6. There can be about 7.7-8.6, 7.7-8.5, 7.7-8.4, 7.7-8.3, 7.7-8.2,7.8-8.6, 7.8-8.5, 7.8-8.4, 7.8-8.3, 7.8-8.2, 7.9-8.6, 7.9-8.5, 7.9-8.4,7.9-8.3, 7.9-8.2, 8.0-8.6, 8.0-8.5, 8.0-8.4, 8.0-8.3, 8.0-8.2, 8.1-8.6,8.1-8.5, 8.1-8.1, 8.1-8.3,or 8.1-8.2 wt % glucose linked only atpositions 1, 3 and 6, in some instances.

A dextran in some aspects can comprise about 0.4, 0.5, 0.6, 0.7, 0.8,0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, or 1.7 wt % glucose linked onlyat (a) positions 1, 2 and 6, or (b) positions 1, 4 and 6. There can beabout 0.4-1.7, 0.4-1.6, 0.4-1.5, 0.4-1.4, 0.4-1.3, 0.5-1.7, 0.5-1.6,0.5-1.5, 0.5-1.4, 0.5-1.3, 0.6-1.7, 0.6-1.6, 0.6-1.5, 0.6-1.4, 0.6-1.3,0.7-1.7, 0.7-1.6, 0.7-1.5, 0.7-1.4, 0.7-1.3, 0.8-1.7, 0.8-1.6, 0.8-1.5,0.8-1.4, 0.8-1.3 wt % glucose linked only at (a) positions 1, 2 and 6,or (b) positions 1, 4 and 6, in some instances.

The glucosidic linkage profile of dextran can be determined usingdextran produced following any protocol disclosed herein. An example ofa suitable linkage determination protocol can be similar to, or the sameas, the protocol disclosed in Example 9: For example, an 0768 gtf enzymereaction that has been deactivated by heating the reaction at about70-90° C. (e.g., 80° C.) for about 5-30 minutes (e.g., 10 minutes) isplaced into dialysis tubing (e.g., made with regenerated cellulose) withan MWCO of 12-14 kDa (e.g., Spectra/Por® 4 Dialysis Tubing, Part No.132706, Spectrum Laboratories, Inc.). The deactivated reaction is thendialyzed against a large volume of water (e.g., 3-5 L) at about 20-25°C. (room temp) over about 4-10 days (e.g., 7 days); this water can beexchanged every day during the dialysis. The dextran product is then (i)precipitated by mixing the dialyzed deactivated reaction with about 1-2×(1.5×) reaction volume of 100% methanol, (ii) washed at least two timeswith the same volume of 100% methanol, and (iii) dried at about 40-50°C. (e.g., 45° C.) (optionally under a vacuum). A dissolvable amount ofdry dextran is dissolved in dimethyl sulfoxide (DMSO) or DMSO/5% LiCl,after which all free hydroxyl groups are methylated (e.g., by sequentialaddition of a NaOH/DMSO slurry followed with iodomethane). Themethylated dextran is then extracted (e.g., into methylene chloride) andhydrolyzed to monomeric units using aqueous trifluoroacetic acid (TFA)at about 110-125° C. (e.g., 120° C.). The TFA is then evaporated andreductive ring opening is done using sodium borodeuteride. The hydroxylgroups created by hydrolyzing the glycosidic linkages are thenacetylated by treating with acetyl chloride and TFA at a temperature ofabout 40-60° C. (e.g., 50° C.). Next, the derivatizing reagents areevaporated and the resulting methylated/acetylated monomers arereconstituted in acetonitrile; this preparation is then analyzed byGC/MS using an appropriate column (e.g., biscyanopropylcyanopropylphenyl polysiloxane). The relative positioning of the methyland acetyl functionalities render species with distinctive retentiontime indices and mass spectra that can be compared to publisheddatabases. In this way, the derivatives of the monomeric units indicatehow each monomer was originally linked in the dextran polymer.

It is believed that dextran herein may be a branched structure in whichthere are long chains (containing mostly or all alpha-1,6-linkages) thatiteratively branch from each other (e.g., a long chain can be a branchfrom another long chain, which in turn can itself be a branch fromanother long chain, and so on). The branched structure may also compriseshort branches from the long chains; these short chains are believed tomostly comprise alpha-1,3 and -1,4 linkages, for example. Branch pointsin the dextran, whether from a long chain branching from another longchain, or a short chain branching from a long chain, appear to comprisealpha-1,3, -1,4, or -1,2 linkages off of a glucose involved in alpha-1,6linkage. On average, about 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%,29%, 30%, 15-35%, 15-30%, 15-25%, 15-20%, 20-35%, 20-30%, 20-25%,25-35%, or 25-30% of all branch points of dextran in some embodimentsbranch into long chains. Most (>98% or 99%) or all the other branchpoints branch into short chains.

The long chains of a dextran branching structure can be similar inlength in some aspects. By being similar in length, it is meant that thelength (DP) of at least 70%, 75%, 80%, 85%, or 90% of all the longchains in a branching structure is within plus/minus 15% (or 10%, 5%) ofthe mean length of all the long chains of the branching structure. Insome aspects, the mean length (average length) of the long chains isabout 10-50 DP (i.e., 10-50 glucose monomers). For example, the meanindividual length of the long chains can be about 10, 15, 16, 17, 18,19, 20, 21, 22, 23, 24, 25, 30, 35, 40, 45, 50, 10-50, 10-40, 10-30,10-25, 10-20, 15-50, 15-40, 15-30, 15-25, 15-20, 20-50, 20-40, 20-30, or20-25 DP.

Dextran long chains in certain embodiments can comprise substantiallyalpha-1,6-glucosidic linkages and a small amount (less than 2.0%) ofalpha-1,3- and/or alpha-1,4-glucosidic linkages. For example, dextranlong chains can comprise about, or at least about, 98%, 98.25%, 98.5%,98.75%, 99%, 99.25%, 99.5%, 99.75%, or 99.9% alpha-1,6-glucosidiclinkages. A dextran long chain in certain embodiments does not comprisealpha-1,4-glucosidic linkages (i.e., such a long chain has mostlyalpha-1,6 linkages and a small amount of alpha-1,3 linkages).Conversely, a dextran long chain in some embodiments does not comprisealpha-1,3-glucosidic linkages (i.e., such a long chain has mostlyalpha-1,6 linkages and a small amount of alpha-1,4 linkages). Anydextran long chain of the above embodiments may further not comprisealpha-1,2-glucosidic linkages, for example. Still in some aspects, adextran long chain can comprise 100% alpha-1,6-glucosidic linkages(excepting the linkage used by such long chain to branch from anotherchain).

Short chains of a dextran molecule in some aspects are one to threeglucose monomers in length and comprise less than about 5-10% of all theglucose monomers of the dextran polymer. At least about 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, 99%, or all of, short chains herein are1-3 glucose monomers in length. The short chains of a dextran moleculecan comprise less than about 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1%of all the glucose monomers of the dextran molecule, for example.

Short chains of a dextran molecule in some aspects can comprisealpha-1,2-, alpha-1,3-, and/or alpha-1,4-glucosidic linkages. Shortchains, when considered all together (not individually) may comprise (i)all three of these linkages, or (ii) alpha-1,3- and alpha-1,4-glucosidiclinkages, for example. It is believed that short chains of a dextranmolecule herein can be heterogeneous (i.e., showing some variation inlinkage profile) or homogeneous (i.e., sharing similar or same linkageprofile) with respect to the other short chains of the dextran.

Dextran in certain embodiments can have an Mw of about, or at leastabout, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120,125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190,195, or 200 million (or any integer between 50 and 200 million) (or anyrange between two of these values). The Mw of dextran can be about50-200, 60-200, 70-200, 80-200, 90-200, 100-200, 110-200, 120-200,50-180, 60-180, 70-180, 80-180, 90-180, 100-180, 110-180, 120-180,50-160, 60-160, 70-160, 80-160, 90-160, 100-160, 110-160, 120-160,50-140, 60-140, 70-140, 80-140, 90-140, 100-140, 110-140, 120-140,50-120, 60-120, 70-120, 80-120, 90-120, 100-120, 110-120, 50-110,60-110, 70-110, 80-110, 90-110, 100-110, 50-100, 60-100, 70-100, 80-100,90-100, or 95-105 million, for example. Any of these Mw's can berepresented in DPw, if desired, by dividing Mw by 162.14.

The z-average radius of gyration of a dextran herein can be about200-280 nm. For example, the z-average Rg can be about 200, 205, 210,215, 220, 225, 230, 235, 240, 245, 250, 255, 260, 265, 270, 275, or 280nm (or any integer between 200-280 nm). As other examples, the z-averageRg can be about 200-280, 200-270, 200-260, 200-250, 200-240, 200-230,220-280, 220-270, 220-260, 220-250, 220-240, 220-230, 230-280, 230-270,230-260, 230-250, 230-240, 240-280, 240-270, 240-260, 240-250, 250-280,250-270, or 250-260 nm.

The Mw and/or z-average Rg of dextran in some aspects can be measuredfollowing a protocol similar to, or the same as, the protocol disclosedin Example 9. For example, a Mw and/or z-average Rg herein can bemeasured by first dissolving dextran produced by an 0768 gtf at 0.4-0.6mg/mL (e.g., ˜0.5 mg/mL) in 0.05-1.0 M (e.g., ˜0.075 M)Tris(hydroxymethyl)aminomethane buffer with 150-250 ppm (e.g., ˜200 ppm)NaN₃. Solvation of dry dextran can be achieved by shaking for 12-18hours at 45-55° C. (e.g., ˜50° C.). The resulting dextran solution canbe entered into a suitable flow injection chromatographic apparatuscomprising a separation module (e.g., Alliance™ 2695 separation modulefrom Waters Corporation, Milford, Mass.) coupled with three onlinedetectors: a differential refractometer (e.g., Waters 2414 refractiveindex detector), a multiangle light scattering (MALS) photometer (e.g.,Heleos™-2 18-angle multiangle MALS photometer) equipped with aquasielastic light scatering (QELS) detector (e.g., QELS detector fromWyatt Technologies, Santa Barbara, Calif.), and a differential capillaryviscometer (e.g., ViscoStar™ differential capillary viscometer fromWyatt). Two suitable size-exclusion columns (e.g., AQUAGEL-OH GUARDcolumns from Agilent Technologies, Santa Clara, Calif.) can be used toseparate the dextran polymer peak from the injection peak, where themobile phase can be the same as the sample solvent (above), the flowrate can be about 0.2 mL/min, the injection volumes can be about 0.1 mL,and column temperature can be about 30° C. Suitable software can be usedfor data acquisition (e.g., Empower™ version 3 software from Waters) andfor multidetector data reduction (Astra™ version 6 software from Wyatt).MALS data can provide weight-average molecular weight (Mw) and z-averageradius of gyration (Rg), and QELS data can provide z-averagehydrodynamic radius, for example.

A dextran herein can be a product of a glucosyltransferase enzymecomprising, or consisting of, an amino acid sequence that is 100%identical to, or at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,or 99% identical to, SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:9,SEQ ID NO:13, or SEQ ID NO:17 (and have gtf activity). Non-limitingexamples of a glucosyltransferase enzyme comprising SEQ ID NO:1 (or arelated sequence) include glucosyltransferase enzymes comprising, orconsisting of, an amino acid sequence that is 100% identical to, or atleast 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to,SEQ ID NO:2 (and have gtf activity). Production of dextran can beaccomplished with a gtf reaction as disclosed herein, for example.Dextran as disclosed in the instant detailed description (e.g.,molecular weight, linkage and branching profile) can optionally becharacterized as a product of a glucosyltransferase enzyme comprising orconsisting of SEQ ID NO:1 or 2 (or a related sequence thereof that is atleast 90% identical [above]). In some other embodiments, aglucosyltransferase enzyme comprises or consists of an amino acidsequence that is 100% identical to, or at least 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, or 99% identical to, the secreted portion (i.e.,signal peptide removed) of the amino acid sequence encoded by SEQ IDNO:6, 10, 14, or 18.

A glucosyltransferase enzyme herein may be from various microbialsources, such as a bacteria or fungus. Examples of bacterialglucosyltransferase enzymes are those derived from a Streptococcusspecies, Leuconostoc species, Lactobacillus species, or Weissellaspecies. Examples of Streptococcus species include S. sobrinus, S.downei, S. salivarius, S. dentirousetti, S. mutans, S. oralis, S.gallolyticus and S. sanguinis. Examples of Leuconostoc species includeL. pseudomesenteroides, L. mesenteroides, L. amelibiosum, L. argentinum,L. carnosum, L. citreum, L. cremoris, L. dextranicum and L. fructosum.Examples of Lactobacillus species include L. fermentum, L. acidophilus,L. delbrueckii, L. helveticus, L. salivarius, L. casei, L. curvatus, L.plantarum, L. sakei, L. brevis, L. buchneri and L. reuteri. Examples ofWeissella species include W. cibaria, W. confusa, W. halotolerans, W.hellenica, W. kandleri, W. kimchii, W. koreensis, W. minor, W.paramesenteroides, W. soli and W. thailandensis. A glucosyltransferasein some aspects is not from L. mesenteroides.

Examples of glucosyltransferase enzymes herein can be any of the aminoacid sequences disclosed herein and that further include 1-300 (or anyinteger there between [e.g., 10, 15, 20, 25, 30, 35, 40, 45, or 50])residues on the N-terminus and/or C-terminus. Such additional residuesmay be from a corresponding wild type sequence from which theglucosyltransferase enzyme is derived, or may be a heterologous sequencesuch as an epitope tag (at either N- or C-terminus) or a heterologoussignal peptide (at N-terminus), for example.

A glucosyltransferase enzyme used to produce dextran herein is typicallyin a mature form lacking an N-terminal signal peptide. An expressionsystem for producing a mature glucosyltransferase enzyme herein mayemploy an enzyme-encoding polynucleotide that further comprises sequenceencoding an N-terminal signal peptide to direct extra-cellularsecretion. The signal peptide in such embodiments is cleaved from theenzyme during the secretion process. The signal peptide may either benative or heterologous to the glucosyltransferase. An example of asignal peptide useful herein is one from a bacterial (e.g., a Bacillusspecies such as B. subtilis) or fungal species. An example of abacterial signal peptide is an aprE signal peptide, such as one fromBacillus (e.g., B. subtilis, see Vogtentanz et al., Protein Expr. Purif.55:40-52, which is incorporated herein by reference).

SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:9, SEQ ID NO:13 and SEQ ID NO:17 areexamples of mature glucosyltransferase enzymes that lack an N-terminalsignal peptide. Since these and related amino acid sequences do notbegin with a methionine residue, it would be understood that anN-terminal start-methionine is preferably added to the sequence(directly or via an intervening heterologous amino acid sequence such asan epitope) if expressing any of these enzymes without using a signalpeptide (such as with an expression system where the enzyme is expressedintracellularly and obtained from a cell lysate).

A glucosyltransferase enzyme in certain embodiments can be produced byany means known in the art. For example, a glucosyltransferase enzymecan be produced recombinantly in a heterologous expression system, suchas a microbial heterologous expression system. Examples of heterologousexpression systems include bacterial (e.g., E. coli such as TOP10,MG1655, or BL21 DE3; Bacillus sp. such as B. subtilis) and eukaryotic(e.g., yeasts such as Pichia sp. and Saccharomyces sp.) expressionsystems.

A glucosyltransferase enzyme disclosed herein may be used in anypurification state (e.g., pure or non-pure). For example, theglucosyltransferase enzyme may be purified and/or isolated prior to itsuse. Examples of glucosyltransferase enzymes that are non-pure includethose in the form of a cell lysate. A cell lysate or extract may beprepared from a bacteria (e.g., E. coli) used to heterologously expressthe enzyme. For example, the bacteria may be subjected to disruptionusing a French pressure cell. In alternative embodiments, bacteria maybe homogenized with a homogenizer (e.g., APV, Rannie, Gaulin). Aglucosyltransferase enzyme is typically soluble in these types ofpreparations. A bacterial cell lysate, extract, or homogenate herein maybe used at about 0.15-0.3% (v/v) in a reaction for producing dextranfrom sucrose.

A heterologous gene expression system for expressing aglucosyltransferase enzyme herein can be designed for protein secretion,for example. A glucosyltransferase enzyme typically comprises a signalpeptide in such embodiments. A glucosyltransferase enzyme in someembodiments does not occur in nature; for example, an enzyme herein isnot believed to be one that is naturally secreted (i.e., mature form)from a microbe (from which the glucosyltransferase enzyme herein couldpossibly have been derived).

The activity of a glucosyltransferase enzyme herein can be determinedusing any method known in the art. For example, glucosyltransferaseenzyme activity can be determined by measuring the production ofreducing sugars (fructose and glucose) in a reaction containing sucrose(˜50 g/L), dextran T10 (˜1 mg/mL) and potassium phosphate buffer (˜pH6.5, 50 mM), where the solution is held at ˜22-25° C. for ˜24-30 hours.The reducing sugars can be measured by adding 0.01 mL of the reaction toa mixture containing ˜1 N NaOH and ˜0.1% triphenyltetrazolium chlorideand then monitoring the increase in absorbance at OD_(480 nm) for ˜fiveminutes. Also for instance, a unit of an enzyme such as gtf 0768(comprising SEQ ID NO:1) herein can be defined as the amount of enzymerequired to consume 1 g of sucrose in 1 hour at 26° C., pH 6.5, and with100 g/L of sucrose.

A dextran as presently disclosed can be a product of aglucosyltransferase as comprised in a glucosyltransferase reaction.

The temperature of a glucosyltransferase reaction herein can becontrolled, if desired. In certain embodiments, the temperature isbetween about 5° C. to about 50° C. The temperature in certain otherembodiments is between about 20° C. to about 40° C. Alternatively, thetemperature may be about 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31,32, 33, 34, 35, 36, 37, 38, 39, or 40° C. The temperature of aglucosyltransferase reaction herein may be maintained using variousmeans known in the art. For example, the temperature can be maintainedby placing the vessel containing the reaction in an air or water bathincubator set at the desired temperature.

The initial concentration of sucrose in a glucosyltransferase reactionherein can be about 20 g/L to 900 g/L, 20 g/L to 400 g/L, 75 g/L to 175g/L, or 50 g/L to 150 g/L. The initial concentration of sucrose can beabout 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160,200, 300, 400, 500, 600, 700, 800, 900, 50-150, 75-125, 90-110, 50-500,100-500, 200-500, 300-500, 400-500, 50-400, 100-400, 200-400, 300-400,50-300, 100-300, 200-300, 50-200, 100-200, or 50-100 g/L (or any integerbetween 20 and 900 g/L), for example. “Initial concentration of sucrose”refers to the sucrose concentration in a gtf reaction just after all thereaction components have been added (at least water, sucrose,glucosyltransferase enzyme).

Sucrose used in a glucosyltransferase reaction herein can be highly pure(≥99.5%) or be of any other purity or grade. For example, sucrose canhave a purity of at least 99.0%, or can be reagent grade sucrose. Asanother example, incompletely refined sucrose can be used. Incompletelyrefined sucrose herein refers to sucrose that has not been processed towhite refined sucrose. Thus, incompletely refined sucrose can becompletely unrefined or partially refined. Examples of unrefined sucroseare “raw sucrose” (“raw sugar”) and solutions thereof. Examples ofpartially refined sucrose have not gone through one, two, three, or morecrystallization steps. The ICUMSA (International Commission for UniformMethods of Sugar Analysis) of incompletely refined sucrose herein can begreater than 150, for example. Sucrose herein may be derived from anyrenewable sugar source such as sugar cane, sugar beets, cassava, sweetsorghum, or corn. Suitable forms of sucrose useful herein arecrystalline form or non-crystalline form (e.g., syrup, cane juice, beetjuice), for example. Additional suitable forms of incompletely refinedsucrose are disclosed in U.S. Appl. Publ. No. 2015/0275256, which isincorporated herein by reference.

Methods of determining ICUMSA values for sucrose are well known in theart and disclosed by the International Commission for Uniform Methods ofSugar Analysis in ICUMSA Methods of Sugar Analysis: Official andTentative Methods Recommended by the International Commission forUniform Methods of Sugar Analysis (ICUMSA) (Ed. H. C. S. de Whalley,Elsevier Pub. Co., 1964), for example, which is incorporated herein byreference. ICUMSA can be measured, for example, by ICUMSA Method GS1/3-7as described by R. J. McCowage, R. M. Urquhart and M. L. Burge(Determination of the Solution Colour of Raw Sugars, Brown Sugars andColoured Syrups at pH 7.0—Official, Verlag Dr Albert Bartens, 2011revision), which is incorporated herein by reference.

The pH of a glucosyltransferase reaction in certain embodiments can bebetween about 4.0 to about 8.0. Alternatively, the pH can be about 4.0,4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, or 8.0. The pH can be adjusted orcontrolled by the addition or incorporation of a suitable buffer,including but not limited to: phosphate, tris, citrate, or a combinationthereof. Buffer concentration in a gtf reaction can be from 0 mM toabout 100 mM, or about 10, 20, or 50 mM, for example.

A glucosyltransferase reaction can be contained within any vesselsuitable for applying one or more of the reaction conditions disclosedherein. For example, a glucosyltransferase reaction herein may be in astainless steel, plastic, or glass vessel or container of a sizesuitable to contain a particular reaction. Such a vessel can optionallybe equipped with a stirring device.

A glucosyltransferase reaction herein can optionally be agitated viastirring or orbital shaking, for example. Such agitation can be at about50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 50-150, 60-140,70-130, 80-120, or 90-110 rpm, for example.

The concentration of glucosyltransferase enzyme in a reaction can be atleast about 15, 20, 25, 30, 35, or 40 U/L, for example. In someembodiments, 15-35, 15-30, 15-25, 20-35, 20-30, 20-25, 25-35, 25-30, or30-35 U/L of glucosyltransferase can be used.

A glucosyltransferase reaction herein can take about 2, 3, 4, 5, 6, 7,8, 9, 10, 12, 18, 24, 30, 36, 48, 60, 72, 84, 96, 18-30, 20-28, or 22-26hours to complete. Reaction time may depend, for example, on certainparameters such as the amount of sucrose and glucosyltransferase enzymeused in the reaction.

All the features herein defining a glucosyltransferase reaction can becombined, accordingly. Simply as an example, a reaction using an 0768glucosyltransferase (comprising SEQ ID NO:1 or related sequence thereof)can initially contain 90-110 g/L (e.g., ˜100 g/L) sucrose, 10-30 mM(e.g., ˜20 mM) sodium phosphate buffer at pH 6.0-7.0 (e.g., ˜pH 6.5),and 20-30 U/L (e.g., ˜25 U/L) enzyme. Such a reaction can be held forabout 20-28 hours (e.g., ˜24 hours) with 50-150 rpm (e.g., ˜100 rpm)shaking at 24-28° C. (e.g., ˜26° C.).

In some embodiments, a glucosyltransferase reaction comprising a gtf0768 enzyme (SEQ ID NO:1 or related sequences) and any amount of sucrosedisclosed herein can be complete (e.g., 95% or more initially providedsucrose depleted) in less than about 24, 22, 20, 18, or 16 hours afterinitiating the reaction. Depletion of sucrose in such a reaction can beabout, or at least about, 3, 4, 5, 6, 7, 8, 9, or 10 times faster than asame or similar reaction, but which comprises a Leuconostocmesenteroides dextran sucrase instead of a gtf 0768 enzyme, for example.

A composition comprising a dextran herein can be non-aqueous (e.g., adry composition). Examples of such embodiments include powders,granules, microcapsules, flakes, or any other form of particulatematter. Other examples include larger compositions such as pellets,bars, kernels, beads, tablets, sticks, or other agglomerates. Anon-aqueous or dry composition herein typically has less than 3, 2, 1,0.5, or 0.1 wt % water comprised therein. The amount of dextran hereinin a non-aqueous or dry composition can be about, or at least about, 1,2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21,22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39,40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 55, 56, 57, 58,59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76,77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94,95, 96, 97, 98, 99, 99.5, or 99.9 wt %, for example. A non-aqueouscomposition herein can be in the form of a household product, personalcare product, pharmaceutical product, industrial product, or foodproduct, for example.

In certain embodiments of the present disclosure, a compositioncomprising a dextran can be an aqueous composition having a viscosity ofabout, or at least about, 25 cPs. Alternatively, an aqueous compositionherein can have a viscosity of about, or at least about, 25, 50, 75,100, 250, 500, 750, 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000,9000, 10000, 11000, 12000, 13000, 14000, 15000, 16000, 17000, 18000,19000, 20000, 25000, 30000, 35000, 40000, 45000, or 50000 cPs (or anyinteger between 25 and 50000 cPs), for example. Examples of aqueouscompositions include hydrocolloids and aqueous solutions.

Viscosity can be measured with an aqueous composition herein at anytemperature between about 3° C. to about 110° C. (or any integer between3 and 110° C.). Alternatively, viscosity can be measured at atemperature between about 4° C. to 30° C., or about 20° C. to 25° C.,for example. Viscosity can be measured at atmospheric pressure (about760 torr) or any other higher or lower pressure.

The viscosity of an aqueous composition disclosed herein can be measuredusing a viscometer or rheometer, or using any other means known in theart. It would be understood by those skilled in the art that aviscometer or rheometer can be used to measure the viscosity of aqueouscompositions herein that exhibits rheological behavior (i.e., havingviscosities that vary with flow conditions). The viscosity of suchembodiments can be measured at a rotational shear rate of about 0.1 to1000 rpm (revolutions per minute), for example. Alternatively, viscositycan be measured at a rotational shear rate of about 10, 60, 150, 250, or600 rpm.

In certain embodiments, viscosity can be measured with an aqueouscomposition in which the constituent dextran was synthesized. Forexample, viscosity can be measured for a gtf reaction herein that is ator near completion. Viscosity can thus be measured with an aqueouscomposition in which the constituent dextran is not purified (e.g.,other components in the composition, aside from water, are present atgreater than 1, 5, or 10 wt %); such a composition can contain one ormore salts, buffers, proteins (e.g., gtf enzymes), sugars (e.g.,fructose, glucose, leucrose, oligosaccharides).

The pH of an aqueous composition disclosed herein can be between about2.0 to about 12.0, for example. Alternatively, pH can be about 2.0, 3.0,4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10.0, 11.0, 12.0; or between 5.0 to about12.0; or between about 4.0 and 8.0; or between about 5.0 and 8.0, forexample.

An aqueous composition herein such as a hydrocolloid or aqueous solutioncan comprise a solvent having about, or at least about, 10 wt % water.In other embodiments, a solvent is about, or at least about, 20, 30, 40,50, 60, 70, 80, 90, or 100 wt % water (or any integer value between 10and 100 wt %), for example.

A dextran herein can be present in an aqueous composition at a wt % ofabout, or at least about, 0.01, 0.05, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7,0.8, 0.9, 1.0, 1.2, 1.4, 1.6, 1.8, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5, 6,7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25,26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43,44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 55, 56, 57, 58, 59, 60, 61, 62,63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80,81, 82, 83, 84, 85, 86, 87, 88, 89, or 90 wt %, for example. Example 8below demonstrates that dextran in certain aspects provides highviscosity to aqueous solutions at relatively low concentrations of thedextran. Thus, certain embodiments of the present disclosure are drawnto aqueous compositions with less than about 30, 29, 28, 27, 26, 25, 24,23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5,4, 3, 2, 1, or 0.5 wt % dextran herein.

An aqueous composition herein can comprise other components in additionto dextran. For example, an aqueous composition can comprise one or moresalts such as a sodium salt (e.g., NaCl, Na₂SO₄). Other non-limitingexamples of salts include those having (i) an aluminum, ammonium,barium, calcium, chromium (II or III), copper (I or II), iron (II orIII), hydrogen, lead (II), lithium, magnesium, manganese (II or III),mercury (I or II), potassium, silver, sodium strontium, tin (II or IV),or zinc cation, and (ii) an acetate, borate, bromate, bromide,carbonate, chlorate, chloride, chlorite, chromate, cyanamide, cyanide,dichromate, dihydrogen phosphate, ferricyanide, ferrocyanide, fluoride,hydrogen carbonate, hydrogen phosphate, hydrogen sulfate, hydrogensulfide, hydrogen sulfite, hydride, hydroxide, hypochlorite, iodate,iodide, nitrate, nitride, nitrite, oxalate, oxide, perchlorate,permanganate, peroxide, phosphate, phosphide, phosphite, silicate,stannate, stannite, sulfate, sulfide, sulfite, tartrate, or thiocyanateanion. Thus, any salt having a cation from (i) above and an anion from(ii) above can be in an aqueous composition, for example. A salt can bepresent in an aqueous composition herein at a wt % of about 0.01 toabout 10.00 (or any hundredth increment between 0.01 and 10.00), forexample.

A composition herein may optionally contain one or more active enzymes.Non-limiting examples of suitable enzymes include proteases, cellulases,hemicellulases, peroxidases, lipolytic enzymes (e.g., metallolipolyticenzymes), xylanases, lipases, phospholipases, esterases (e.g.,arylesterase, polyesterase), perhydrolases, cutinases, pectinases,pectate lyases, mannanases, keratinases, reductases, oxidases (e.g.,choline oxidase), phenoloxidases, lipoxygenases, ligninases,pullulanases, tannases, pentosanases, malanases, beta-glucanases,arabinosidases, hyaluronidases, chondroitinases, laccases,metalloproteinases, amadoriases, glucoamylases, arabinofuranosidases,phytases, isomerases, transferases and amylases. If an enzyme(s) isincluded, it may be comprised in a composition herein at about0.0001-0.1 wt % (e.g., 0.01-0.03 wt %) active enzyme (e.g., calculatedas pure enzyme protein), for example.

A cellulase herein can have endocellulase activity (EC 3.2.1.4),exocellulase activity (EC 3.2.1.91), or cellobiase activity (EC3.2.1.21). A cellulase herein is an “active cellulase” having activityunder suitable conditions for maintaining cellulase activity; it iswithin the skill of the art to determine such suitable conditions.

A cellulase herein may be derived from any microbial source, such as abacteria or fungus. Chemically-modified cellulases or protein-engineeredmutant cellulases are included. Suitable cellulases include, but are notlimited to, cellulases from the genera Bacillus, Pseudomonas,Streptomyces, Trichoderma, Humicola, Fusarium, Thielavia and Acremonium.As other examples, a cellulase may be derived from Humicola insolens,Myceliophthora thermophila or Fusarium oxysporum; these and othercellulases are disclosed in U.S. Pat. Nos. 4,435,307, 5,648,263,5,691,178, 5,776,757 and 7,604,974, which are all incorporated herein byreference. Exemplary Trichoderma reesei cellulases are disclosed in U.S.Pat. Nos. 4,689,297, 5,814,501, 5,324,649, and International PatentAppl. Publ. Nos. WO92/06221 and WO92/06165, all of which areincorporated herein by reference. Exemplary Bacillus cellulases aredisclosed in U.S. Pat. No. 6,562,612, which is incorporated herein byreference. A cellulase, such as any of the foregoing, preferably is in amature form lacking an N-terminal signal peptide. Commercially availablecellulases useful herein include CELLUZYME® and CAREZYME® (NovozymesA/S); CLAZINASE® and PURADAX® HA (DuPont Industrial Biosciences), andKAC-500(B)® (Kao Corporation).

Alternatively, a cellulase herein may be produced by any means known inthe art, such as described in U.S. Pat. Nos. 4,435,307, 5,776,757 and7,604,974, which are incorporated herein by reference. For example, acellulase may be produced recombinantly in a heterologous expressionsystem, such as a microbial or fungal heterologous expression system.Examples of heterologous expression systems include bacterial (e.g., E.coli, Bacillus sp.) and eukaryotic systems. Eukaryotic systems canemploy yeast (e.g., Pichia sp., Saccharomyces sp.) or fungal (e.g.,Trichoderma sp. such as T. reesei, Aspergillus species such as A. niger)expression systems, for example.

One or more cellulases can be directly added as an ingredient whenpreparing a composition disclosed herein. Alternatively, one or morecellulases can be indirectly (inadvertently) provided in the disclosedcomposition. For example, cellulase can be provided in a compositionherein by virtue of being present in a non-cellulase enzyme preparationused for preparing a composition. Cellulase in compositions in whichcellulase is indirectly provided thereto can be present at about 0.1-10ppb (e.g., less than 1 ppm), for example. A contemplated benefit of acomposition herein, by virtue of employing a dextran compound, is thatnon-cellulase enzyme preparations that might have background cellulaseactivity can be used without concern that the desired effects of thedextran will be negated by the background cellulase activity.

A cellulase in certain embodiments can be thermostable. Cellulasethermostability refers to the ability of the enzyme to retain activityafter exposure to an elevated temperature (e.g. about 60-70° C.) for aperiod of time (e.g., about 30-60 minutes). The thermostability of acellulase can be measured by its half-life (t½) given in minutes, hours,or days, during which time period half the cellulase activity is lostunder defined conditions.

A cellulase in certain embodiments can be stable to a wide range of pHvalues (e.g. neutral or alkaline pH such as pH of ˜7.0 to ˜11.0). Suchenzymes can remain stable for a predetermined period of time (e.g., atleast about 15 min., 30 min., or 1 hour) under such pH conditions.

At least one, two, or more cellulases may be included in thecomposition. The total amount of cellulase in a composition hereintypically is an amount that is suitable for the purpose of usingcellulase in the composition (an “effective amount”). For example, aneffective amount of cellulase in a composition intended for improvingthe feel and/or appearance of a cellulose-containing fabric is an amountthat produces measurable improvements in the feel of the fabric (e.g.,improving fabric smoothness and/or appearance, removing pills andfibrils which tend to reduce fabric appearance sharpness). As anotherexample, an effective amount of cellulase in a fabric stonewashingcomposition herein is that amount which will provide the desired effect(e.g., to produce a worn and faded look in seams and on fabric panels).The amount of cellulase in a composition herein can also depend on theprocess parameters in which the composition is employed (e.g.,equipment, temperature, time, and the like) and cellulase activity, forexample. The effective concentration of cellulase in an aqueouscomposition in which a fabric is treated can be readily determined by askilled artisan. In fabric care processes, cellulase can be present inan aqueous composition (e.g., wash liquor) in which a fabric is treatedin a concentration that is minimally about 0.01-0.1 ppm total cellulaseprotein, or about 0.1-10 ppb total cellulase protein (e.g., less than 1ppm), to maximally about 100, 200, 500, 1000, 2000, 3000, 4000, or 5000ppm total cellulase protein, for example.

Dextran polymers provided herein are believed to be mostly or completelystable (resistant) to being degraded by cellulase. For example, thepercent degradation of a dextran herein by one or more cellulases isbelieved to be less than 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1%, oris 0%. Such percent degradation can be determined, for example, bycomparing the molecular weight of dextran polymer before and aftertreatment with a cellulase for a period of time (e.g., ˜24 hours).

Aqueous compositions in certain embodiments are believed to have shearthinning behavior or shear thickening behavior. Shear thinning behavioris observed as a decrease in viscosity of the aqueous composition asshear rate increases, whereas shear thickening behavior is observed asan increase in viscosity of the aqueous composition as shear rateincreases. Modification of the shear thinning behavior or shearthickening behavior of an aqueous composition herein can be due to theadmixture of a dextran to the aqueous composition. Thus, one or moredextran compounds of the present disclosure can be added to an aqueouscomposition to modify its rheological profile (i.e., the flow propertiesof an aqueous liquid, solution, or mixture are modified). Also, one ormore dextran compounds can be added to an aqueous composition to modifyits viscosity.

The rheological properties of aqueous compositions herein can beobserved by measuring viscosity over an increasing rotational shear rate(e.g., from about 0.1 rpm to about 1000 rpm). For example, shearthinning behavior of an aqueous composition disclosed herein can beobserved as a decrease in viscosity (cPs) by about, or at least about,5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%,75%, 80%, 85%, 90%, or 95% (or any integer between 5% and 95%) as therotational shear rate increases from about 10 rpm to 60 rpm, 10 rpm to150 rpm, 10 rpm to 250 rpm, 60 rpm to 150 rpm, 60 rpm to 250 rpm, or 150rpm to 250 rpm. As another example, shear thickening behavior of anaqueous composition disclosed herein can be observed as an increase inviscosity (cPs) by about, or at least about, 5%, 10%, 15%, 20%, 25%,30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%,100%, 125%, 150%, 175%, or 200% (or any integer between 5% and 200%) asthe rotational shear rate increases from about 10 rpm to 60 rpm, 10 rpmto 150 rpm, 10 rpm to 250 rpm, 60 rpm to 150 rpm, 60 rpm to 250 rpm, or150 rpm to 250 rpm.

An aqueous composition disclosed herein can be in the form of, and/orcomprised in, a food product, personal care product, pharmaceuticalproduct, household product, or industrial product, such as any of thoseproducts described below. Dextran compounds herein can be used asthickening agents in each of these products. Such a thickening agent maybe used in conjunction with one or more other types of thickening agentsif desired, such as those disclosed in U.S. Pat. No. 8,541,041, thedisclosure of which is incorporated herein by reference in its entirety.

Dextran compounds disclosed herein are believed to be useful forproviding one or more of the following physical properties to a personalcare product, pharmaceutical product, household product, industrialproduct, or food product: thickening, freeze/thaw stability, lubricity,moisture retention and release, texture, consistency, shape retention,emulsification, binding, suspension, dispersion, gelation, reducedmineral hardness, for example. Examples of a concentration or amount ofa dextran in a product can be any of the weight percentages providedherein, for example.

Personal care products herein are not particularly limited and include,for example, skin care compositions, cosmetic compositions, antifungalcompositions, and antibacterial compositions. Personal care productsherein may be in the form of, for example, lotions, creams, pastes,balms, ointments, pomades, gels, liquids, combinations of these and thelike. The personal care products disclosed herein can include at leastone active ingredient, if desired. An active ingredient is generallyrecognized as an ingredient that causes an intended pharmacologicaleffect.

In certain embodiments, a skin care product can be applied to skin foraddressing skin damage related to a lack of moisture. A skin careproduct may also be used to address the visual appearance of skin (e.g.,reduce the appearance of flaky, cracked, and/or red skin) and/or thetactile feel of the skin (e.g., reduce roughness and/or dryness of theskin while improved the softness and subtleness of the skin). A skincare product typically may include at least one active ingredient forthe treatment or prevention of skin ailments, providing a cosmeticeffect, or for providing a moisturizing benefit to skin, such as zincoxide, petrolatum, white petrolatum, mineral oil, cod liver oil,lanolin, dimethicone, hard fat, vitamin A, allantoin, calamine, kaolin,glycerin, or colloidal oatmeal, and combinations of these. A skin careproduct may include one or more natural moisturizing factors such asceramides, hyaluronic acid, glycerin, squalane, amino acids,cholesterol, fatty acids, triglycerides, phospholipids,glycosphingolipids, urea, linoleic acid, glycosaminoglycans,mucopolysaccharide, sodium lactate, or sodium pyrrolidone carboxylate,for example. Other ingredients that may be included in a skin careproduct include, without limitation, glycerides, apricot kernel oil,canola oil, squalane, squalene, coconut oil, corn oil, jojoba oil,jojoba wax, lecithin, olive oil, safflower oil, sesame oil, shea butter,soybean oil, sweet almond oil, sunflower oil, tea tree oil, shea butter,palm oil, cholesterol, cholesterol esters, wax esters, fatty acids, andorange oil.

A personal care product herein can also be in the form of makeup,lipstick, mascara, rouge, foundation, blush, eyeliner, lip liner, lipgloss, other cosmetics, sunscreen, sun block, nail polish, nailconditioner, bath gel, shower gel, body wash, face wash, lip balm, skinconditioner, cold cream, moisturizer, body spray, soap, body scrub,exfoliant, astringent, scruffing lotion, depilatory, permanent wavingsolution, antidandruff formulation, antiperspirant composition,deodorant, shaving product, pre-shaving product, after-shaving product,cleanser, skin gel, rinse, dentifrice composition, toothpaste, ormouthwash, for example. An example of a personal care product (e.g., acleanser, soap, scrub, cosmetic) comprises a carrier or exfoliationagent (e.g., jojoba beads [jojoba ester beads]) (e.g., about 1-10, 3-7,4-6, or 5 wt %); such an agent may optionally be dispersed within theproduct.

A personal care product in some aspects can be a hair care product.Examples of hair care products herein include shampoo, hair conditioner(leave-in or rinse-out), cream rinse, hair dye, hair coloring product,hair shine product, hair serum, hair anti-frizz product, hair split-endrepair product, mousse, hair spray, and styling gel. A hair care productcan be in the form of a liquid, paste, gel, solid, or powder in someembodiments. A hair care product as presently disclosed typicallycomprises one or more of the following ingredients, which are generallyused to formulate hair care products: anionic surfactants such aspolyoxyethylenelauryl ether sodium sulfate; cationic surfactants such asstearyltrimethylammonium chloride and/or distearyltrimethylammoniumchloride; nonionic surfactants such as glyceryl monostearate, sorbitanmonopalmitate and/or polyoxyethylenecetyl ether; wetting agents such aspropylene glycol, 1,3-butylene glycol, glycerin, sorbitol, pyroglutamicacid salts, amino acids and/or trimethylglycine; hydrocarbons such asliquid paraffins, petrolatum, solid paraffins, squalane and/or olefinoligomers; higher alcohols such as stearyl alcohol and/or cetyl alcohol;superfatting agents; antidandruff agents; disinfectants;anti-inflammatory agents; crude drugs; water-soluble polymers such asmethyl cellulose, hydroxycellulose and/or partially deacetylated chitin(in addition to one or more dextrans as disclosed herein); antisepticssuch as paraben; ultra-violet light absorbers; pearling agents; pHadjustors; perfumes; and pigments.

A pharmaceutical product herein can be in the form of an emulsion,liquid, elixir, gel, suspension, solution, cream, or ointment, forexample. Also, a pharmaceutical product herein can be in the form of anyof the personal care products disclosed herein, such as an antibacterialor antifungal composition. A pharmaceutical product can further compriseone or more pharmaceutically acceptable carriers, diluents, and/orpharmaceutically acceptable salts. A dextran compound disclosed hereincan also be used in capsules, encapsulants, tablet coatings, and as anexcipients for medicaments and drugs.

Non-limiting examples of food products herein include vegetable, meat,and soy patties; reformed seafood; reformed cheese sticks; cream soups;gravies and sauces; salad dressing; mayonnaise; onion rings; jams,jellies, and syrups; pie filling; potato products such as French friesand extruded fries; batters for fried foods, pancakes/waffles and cakes;pet foods; confectioneries (candy); beverages; frozen desserts; icecream; cultured dairy products such as cottage cheese, yogurt, cheeses,and sour creams; cake icing and glazes; whipped topping; leavened andunleavened baked goods; and the like.

In certain embodiments, dextran herein can be comprised in a foodstuffor any other ingestible material (e.g., enteral pharmaceuticalpreparation) in an amount that provides the desired degree of thickeningand/or dispersion. For example, the concentration or amount of dextranin a product can be about 0.1-3 wt %, 0.1-4 wt %, 0.1-5 wt %, or 0.1-10wt %.

A household and/or industrial product herein can be in the form ofdrywall tape-joint compounds; mortars; grouts; cement plasters; sprayplasters; cement stucco; adhesives; pastes; wall/ceiling texturizers;binders and processing aids for tape casting, extrusion forming,injection molding and ceramics; spray adherents andsuspending/dispersing aids for pesticides, herbicides, and fertilizers;fabric care products such as fabric softeners and laundry detergents;hard surface cleaners; air fresheners; polymer emulsions; gels such aswater-based gels; surfactant solutions; paints such as water-basedpaints; protective coatings; adhesives; sealants and caulks; inks suchas water-based ink; metal-working fluids; or emulsion-based metalcleaning fluids used in electroplating, phosphatizing, galvanizingand/or general metal cleaning operations, for example.

A dextran compound disclosed herein can be comprised in a personal careproduct, pharmaceutical product, household product, or industrialproduct in an amount that provides a desired degree of thickening and/ordispersion, for example. Examples of a concentration or amount of adextran compound in a product can be any of the weight percentagesprovided above, for example.

An aqueous composition in some aspects can comprise about 0.5-2.0 wt %dextran herein (e.g., ˜1.0 wt %), about 15-25 wt % (e.g., ˜20 wt %) ofmoisturizer such as oil (e.g., mineral oil), about 4-6 wt % (˜5 wt %)surfactant/emulsifier (e.g., one or both of sorbitan monooleate orpolysorbate 80, such as ˜2.6 wt % sorbitan monooleate and ˜2.4 wt %polysorbate 80), optionally 0.25-1.0 wt % (e.g., 0.5 wt %) preservative(e.g., preservative comprising one or more of propylene glycol,diazolidinyl urea, methylparaben, or propylparaben [e.g., Germaben®II]), and optionally one or more other ingredients. Such compositionscan be in the form of an emulsion, for example. In these and some otherrelated aspects, dextran as presently disclosed can be used as asubstitute for compounds (e.g., xanthan gum, crosslinked polyacrylicacid polymers such as Carbopol® Ultrez 10) typically used to provideviscosity to certain consumer products such as personal care (e.g.,lotion), food, and/or pharmaceutical products. Still in some aspects thesensory experience rating of an aqueous composition (e.g., personal careitem such as lotion), as measured by ASTM E1490-3 (“Standard Practicefor Descriptive Skinfeel Analysis of Creams and Lotions”, ASTMInternational, West Conshohocken, Pa., 2003, DOI: 10.1520/E1490-03,incorporated herein by reference), can be less than about 8, 7, or 6,where each of rub-out sliminess, afterfeel stickiness, pick-upstringiness and pick-up stickiness are measured in the evaluation.

A food product herein can be in the form of a confectionery, forexample. A confectionary herein can contain one or more sugars (e.g.,sucrose, fructose, dextrose) for sweetening, or otherwise be sugar-free.

Examples of confectioneries herein include boiled sugars (hard boiledcandies [i.e., hard candy]), dragees, jelly candies, gums, licorice,chews, caramels, toffee, fudge, chewing gums, bubble gums, nougat, chewypastes, halawa, tablets, lozenges, icing, frosting, pudding, and gels(e.g., fruit gels, gelatin dessert). Other examples of confectioneriesinclude aerated confectioneries such as marshmallows, and bakedconfectioneries.

A confectionery herein can optionally be prepared with chocolate, in anyform (e.g., bars, candies, bonbons, truffles, lentils). A confectionarycan be coated with chocolate, sugar-coated, candied, glazed, and/orfilm-coated, for example. Film-coating processes typically compriseapplying to the surface of a confectionery a film-forming liquidcomposition which becomes, after drying, a protective film. Thisfilm-coating serves, for example, to protect the active principlescontained in the confectionery; to protect the confectionery itself frommoisture, shocks, and/or friability; and/or to confer the confectioneryattractive visual properties (e.g., shine, uniform color, smoothsurface).

In certain embodiments, a confectionery can be filled with a fillingthat is liquid, pasty, solid, or powdered. Dextran herein can becomprised in such a filling, in which case dextran is optionally alsoincluded in the confectionery component being filled.

A confectionery herein is optionally sugar-free, comprising no sugar andtypically instead having one or more artificial and/or non-sugarsweeteners (optionally non-caloric) (e.g., aspartame, saccharin, STEVIA,SUCRALOSE). A sugar-free confectionery in certain embodiments cancomprise one or more polyols (e.g., erythritol, glycerol, lactitol,mannitol, maltitol, xylitol), soluble fibers, and/or proteins in placeof sugar.

A food product herein can be in the form of a pet food, for example. Apet food herein can be a food for a domesticated animal such as a dog orcat (or any other companion animal), for example. A pet food in certainembodiments provides to a domestic animal one or more of the following:necessary dietary requirements, treats (e.g., dog biscuits), foodsupplements. Examples of pet food include dry pet food (e.g., kernels,kibbles), semi-moist compositions, wet pet food (e.g., canned pet food),or any combination thereof. Wet pet food typically has a moisturecontent over 65%. Semi-moist pet food typically has a moisture contentof 20-65% and can include humectants such as propylene glycol, potassiumsorbate, and ingredients that prevent microbial growth (bacteria andmold). Dry pet food typically has a moisture content less than 20% andits processing usually includes extruding, drying and/or baking. A petfood can optionally be in the form of a gravy, yogurt, powder,suspension, chew, or treat (e.g., biscuits); all these compositions canalso be used as pet food supplements, if desired. Pet treats can besemi-moist chewable treats; dry treats; chewable bones; baked, extrudedor stamped treats; or confection treats, for example. Examples of petfood compositions/formulations in which a dextran herein can be addedinclude those disclosed in U.S. Patent Appl. Publ. Nos. 2013/0280352 and2010/0159103, and U.S. Pat. No. 6,977,084, which are all incorporatedherein by reference.

Compositions disclosed herein can be in the form of a fabric carecomposition. A fabric care composition herein can be used for hand wash,machine wash and/or other purposes such as soaking and/or pretreatmentof fabrics, for example. A fabric care composition may take the form of,for example, a laundry detergent; fabric conditioner; any wash-, rinse-,or dryer-added product; unit dose or spray. Fabric care compositions ina liquid form may be in the form of an aqueous composition as disclosedherein. In other aspects, a fabric care composition can be in a dry formsuch as a granular detergent or dryer-added fabric softener sheet. Othernon-limiting examples of fabric care compositions herein include:granular or powder-form all-purpose or heavy-duty washing agents;liquid, gel or paste-form all-purpose or heavy-duty washing agents;liquid or dry fine-fabric (e.g. delicates) detergents; cleaningauxiliaries such as bleach additives, “stain-stick”, or pre-treatments;substrate-laden products such as dry and wetted wipes, pads, or sponges;sprays and mists.

A detergent composition herein may be in any useful form, e.g., aspowders, granules, pastes, bars, unit dose, or liquid. A liquiddetergent may be aqueous, typically containing up to about 70 wt % ofwater and 0 wt % to about 30 wt % of organic solvent. It may also be inthe form of a compact gel type containing only about 30 wt % water.

A detergent composition herein typically comprises one or moresurfactants, wherein the surfactant is selected from nonionicsurfactants, anionic surfactants, cationic surfactants, ampholyticsurfactants, zwitterionic surfactants, semi-polar nonionic surfactantsand mixtures thereof. In some embodiments, the surfactant is present ata level of from about 0.1% to about 60%, while in alternativeembodiments the level is from about 1% to about 50%, while in stillfurther embodiments the level is from about 5% to about 40%, by weightof the detergent composition. A detergent will usually contain 0 wt % toabout 50 wt % of an anionic surfactant such as linearalkylbenzenesulfonate (LAS), alpha-olefinsulfonate (AOS), alkyl sulfate(fatty alcohol sulfate) (AS), alcohol ethoxysulfate (AEOS or AES),secondary alkanesulfonates (SAS), alpha-sulfo fatty acid methyl esters,alkyl- or alkenylsuccinic acid, or soap. In addition, a detergentcomposition may optionally contain 0 wt % to about 40 wt % of a nonionicsurfactant such as alcohol ethoxylate (AEO or AE), carboxylated alcoholethoxylates, nonylphenol ethoxylate, alkylpolyglycoside,alkyldimethylamineoxide, ethoxylated fatty acid monoethanolamide, fattyacid monoethanolamide, or polyhydroxy alkyl fatty acid amide (asdescribed for example in WO92/06154, which is incorporated herein byreference).

A detergent composition herein typically comprises one or more detergentbuilders or builder systems. One or more oxidized poly alpha-1,3-glucancompounds can be included as a builder, for example. In some aspects,oxidized poly alpha-1,3-glucan can be included as a co-builder, in whichit is used together with one or more additional builders such as anydisclosed herein. Oxidized poly alpha-1,3-glucan compounds for useherein are disclosed in U.S. Patent Appl. Publ. No. 2015/0259439. Insome embodiments incorporating at least one builder, the cleaningcompositions comprise at least about 1%, from about 3% to about 60%, oreven from about 5% to about 40%, builder by weight of the composition.Builders (in addition to oxidized poly alpha-1,3-glucan) include, butare not limited to, alkali metal, ammonium and alkanolammonium salts ofpolyphosphates, alkali metal silicates, alkaline earth and alkali metalcarbonates, aluminosilicates, polycarboxylate compounds, etherhydroxypolycarboxylates, copolymers of maleic anhydride with ethylene orvinyl methyl ether, 1,3,5-trihydroxy benzene-2,4,6-trisulphonic acid,and carboxymethyloxysuccinic acid, various alkali metal, ammonium andsubstituted ammonium salts of polyacetic acids such as ethylenediaminetetraacetic acid and nitrilotriacetic acid, as well as polycarboxylatessuch as mellitic acid, succinic acid, citric acid, oxydisuccinic acid,polymaleic acid, benzene 1,3,5-tricarboxylic acid,carboxymethyloxysuccinic acid, and soluble salts thereof. Indeed, it iscontemplated that any suitable builder will find use in variousembodiments of the present disclosure. Additional examples of adetergent builder or complexing agent include zeolite, diphosphate,triphosphate, phosphonate, citrate, nitrilotriacetic acid (NTA),ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaaceticacid (DTMPA), alkyl- or alkenylsuccinic acid, soluble silicates orlayered silicates (e.g., SKS-6 from Hoechst).

In some embodiments, builders form water-soluble hardness ion complexes(e.g., sequestering builders), such as citrates and polyphosphates(e.g., sodium tripolyphosphate and sodium tripolyphosphate hexahydrate,potassium tripolyphosphate, and mixed sodium and potassiumtripolyphosphate, etc.). It is contemplated that any suitable builderwill find use in the present disclosure, including those known in theart (See, e.g., EP2100949).

In some embodiments, suitable builders can include phosphate buildersand non-phosphate builders. In some embodiments, a builder is aphosphate builder. In some embodiments, a builder is a non-phosphatebuilder. A builder can be used in a level of from 0.1% to 80%, or from5% to 60%, or from 10% to 50%, by weight of the composition. In someembodiments, the product comprises a mixture of phosphate andnon-phosphate builders. Suitable phosphate builders includemono-phosphates, di-phosphates, tri-polyphosphates oroligomeric-polyphosphates, including the alkali metal salts of thesecompounds, including the sodium salts. In some embodiments, a buildercan be sodium tripolyphosphate (STPP). Additionally, the composition cancomprise carbonate and/or citrate, preferably citrate that helps toachieve a neutral pH composition. Other suitable non-phosphate buildersinclude homopolymers and copolymers of polycarboxylic acids and theirpartially or completely neutralized salts, monomeric polycarboxylicacids and hydroxycarboxylic acids and their salts. In some embodiments,salts of the above mentioned compounds include ammonium and/or alkalimetal salts, i.e., lithium, sodium, and potassium salts, includingsodium salts. Suitable polycarboxylic acids include acyclic, alicyclic,hetero-cyclic and aromatic carboxylic acids, wherein in someembodiments, they can contain at least two carboxyl groups which are ineach case separated from one another by, in some instances, no more thantwo carbon atoms.

A detergent composition herein can comprise at least one chelatingagent. Suitable chelating agents include, but are not limited to copper,iron and/or manganese chelating agents and mixtures thereof. Inembodiments in which at least one chelating agent is used, thecomposition comprises from about 0.1% to about 15%, or even from about3.0% to about 10%, chelating agent by weight of the composition.

A detergent composition herein can comprise at least one deposition aid.Suitable deposition aids include, but are not limited to, polyethyleneglycol, polypropylene glycol, polycarboxylate, soil release polymerssuch as polytelephthalic acid, clays such as kaolinite, montmorillonite,atapulgite, illite, bentonite, halloysite, and mixtures thereof.

A detergent composition herein can comprise one or more dye transferinhibiting agents. Suitable polymeric dye transfer inhibiting agentsinclude, but are not limited to, polyvinylpyrrolidone polymers,polyamine N-oxide polymers, copolymers of N-vinylpyrrolidone andN-vinylimidazole, polyvinyloxazolidones and polyvinylimidazoles ormixtures thereof. Additional dye transfer inhibiting agents includemanganese phthalocyanine, peroxidases, polyvinylpyrrolidone polymers,polyamine N-oxide polymers, copolymers of N-vinylpyrrolidone andN-vinylimidazole, polyvinyloxazolidones and polyvinylimidazoles and/ormixtures thereof; chelating agents examples of which includeethylene-diamine-tetraacetic acid (EDTA); diethylene triamine pentamethylene phosphonic acid (DTPMP); hydroxy-ethane diphosphonic acid(HEDP); ethylenediamine N,N′-disuccinic acid (EDDS); methyl glycinediacetic acid (MGDA); diethylene triamine penta acetic acid (DTPA);propylene diamine tetracetic acid (PDT A); 2-hydroxypyridine-N-oxide(HPNO); or methyl glycine diacetic acid (MGDA); glutamic acidN,N-diacetic acid (N,N-dicarboxymethyl glutamic acid tetrasodium salt(GLDA); nitrilotriacetic acid (NTA); 4,5-dihydroxy-m-benzenedisulfonicacid; citric acid and any salts thereof; N-hydroxyethylethylenediaminetri-acetic acid (HEDTA), triethylenetetraaminehexaaceticacid (TTHA), N-hydroxyethyliminodiacetic acid (HEIDA),dihydroxyethylglycine (DHEG), ethylenediaminetetrapropionic acid (EDTP)and derivatives thereof, which can be used alone or in combination withany of the above. In embodiments in which at least one dye transferinhibiting agent is used, a composition herein may comprise from about0.0001% to about 10%, from about 0.01% to about 5%, or even from about0.1% to about 3%, by weight of the composition.

A detergent composition herein can comprise silicates. In some of theseembodiments, sodium silicates (e.g., sodium disilicate, sodiummetasilicate, and/or crystalline phyllosilicates) find use. In someembodiments, silicates are present at a level of from about 1% to about20% by weight of the composition. In some embodiments, silicates arepresent at a level of from about 5% to about 15% by weight of thecomposition.

A detergent composition herein can comprise dispersants. Suitablewater-soluble organic materials include, but are not limited to thehomo- or co-polymeric acids or their salts, in which the polycarboxylicacid comprises at least two carboxyl radicals separated from each otherby not more than two carbon atoms.

A detergent composition herein may additionally comprise one or moreenzymes. Examples of enzymes include proteases, cellulases,hemicellulases, peroxidases, lipolytic enzymes (e.g., metallolipolyticenzymes), xylanases, lipases, phospholipases, esterases (e.g.,arylesterase, polyesterase), perhydrolases, cutinases, pectinases,pectate lyases, mannanases, keratinases, reductases, oxidases (e.g.,choline oxidase, phenoloxidase), phenoloxidases, lipoxygenases,ligninases, pullulanases, tannases, pentosanases, malanases,beta-glucanases, arabinosidases, hyaluronidases, chondroitinases,laccases, metalloproteinases, amadoriases, glucoamylases,alpha-amylases, beta-amylases, galactosidases, galactanases, catalases,carageenases, hyaluronidases, keratinases, lactases, ligninases,peroxidases, phosphatases, polygalacturonases, pullulanases,rhamnogalactouronases, tannases, transglutaminases, xyloglucanases,xylosidases, metalloproteases, arabinofuranosidases, phytases,isomerases, transferases and/or amylases in any combination.

Any cellulase disclosed above is contemplated for use in the discloseddetergent compositions. Suitable cellulases include, but are not limitedto Humicola insolens cellulases (See e.g., U.S. Pat. No. 4,435,307).Exemplary cellulases contemplated for use herein are those having colorcare benefit for a textile. Examples of cellulases that provide a colorcare benefit are disclosed in EP0495257, EP0531372, EP531315,WO96/11262, WO96/29397, WO94/07998; WO98/12307; WO95/24471, WO98/08940,and U.S. Pat. Nos. 5,457,046, 5,686,593 and 5,763,254,all of which areincorporated herein by reference. Examples of commercially availablecellulases useful in a detergent include CELLUSOFT®, CELLUCLEAN®,CELLUZYME®, and CAREZYME® (Novo Nordisk A/S and Novozymes A/S);CLAZINASE®, PURADAX HA®, and REVITALENZ™ (DuPont IndustrialBiosciences); BIOTOUCH® (AB Enzymes); and KAC-500(B)™ (Kao Corporation).Additional cellulases are disclosed in, e.g., U.S. Pat. No. 7,595,182,U.S. Pat. No. 8,569,033, U.S. Pat. No. 7,138,263, U.S. Pat. No.3,844,890, U.S. Pat. No. 4,435,307, U.S. Pat. No. 4,435,307, andGB2095275.

In some embodiments, a detergent composition can comprise one or moreenzymes (e.g., any disclosed herein), each at a level from about0.00001% to about 10% by weight of the composition and the balance ofcleaning adjunct materials by weight of composition. In some otherembodiments, a detergent composition can also comprise each enzyme at alevel of about 0.0001% to about 10%, about 0.001% to about 5%, about0.001% to about 2%, or about 0.005% to about 0.5%, by weight of thecomposition.

Suitable proteases include those of animal, vegetable or microbialorigin. In some embodiments, microbial proteases are used. In someembodiments, chemically or genetically modified mutants are included. Insome embodiments, the protease is a serine protease, preferably analkaline microbial protease or a trypsin-like protease. Examples ofalkaline proteases include subtilisins, especially those derived fromBacillus (e.g., subtilisin, lentus, amyloliquefaciens, subtilisinCarlsberg, subtilisin 309, subtilisin 147 and subtilisin 168).Additional examples include those mutant proteases described in U.S.Pat. Nos. RE34606, 5,955,340, 5,700,676, 6,312,936 and 6,482,628, all ofwhich are incorporated herein by reference. Additional protease examplesinclude, but are not limited to, trypsin (e.g., of porcine or bovineorigin), and the Fusarium protease described in WO89/06270. In someembodiments, commercially available protease enzymes include, but arenot limited to, MAXATASE®, MAXACAL™, MAXAPEM™, OPTICLEAN®, OPTIMASE®,PROPERASE®, PURAFECT®, PURAFECT® OXP, PURAMAX™, EXCELLASE™, PREFERENZ™proteases (e.g. P100, P110, P280), EFFECTENZ™ proteases (e.g. P1000,P1050, P2000), EXCELLENZ™ proteases (e.g. P1000), ULTIMASE®, andPURAFAST™ (Genencor); ALCALASE®, SAVINASE®, PRIMASE®, DURAZYM™,POLARZYME®, OVOZYME®, KANNASE®, LIQUANASE®, NEUTRASE®, RELASE® andESPERASE® (Novozymes); BLAP™ and BLAP™ variants (HenkelKommanditgesellschaft auf Aktien, Duesseldorf, Germany), and KAP (B.alkalophilus subtilisin; Kao Corp., Tokyo, Japan). Various proteases aredescribed in WO95/23221, WO92/21760, WO09/149200, WO09/149144,WO09/149145, WO11/072099, WO10/056640, WO10/056653, WO11/140364,WO12/151534, U.S. Pat. Publ. No. 2008/0090747, and U.S. Pat. Nos.5,801,039, 5,340,735, 5,500,364, 5,855,625, RE34606, 5,955,340,5,700,676, 6,312,936, 6,482,628, 8,530,219, and various other patents.In some further embodiments, neutral metalloproteases find use in thepresent disclosure, including but not limited to, the neutralmetalloproteases described in WO1999014341, WO1999033960, WO1999014342,WO1999034003, WO2007044993, WO2009058303 and WO2009058661, all of whichare incorporated herein by reference. Exemplary metalloproteases includenprE, the recombinant form of neutral metalloprotease expressed inBacillus subtilis (See e.g., WO07/044993), and PMN, the purified neutralmetalloprotease from Bacillus amyloliquefaciens.

Suitable mannanases include, but are not limited to, those of bacterialor fungal origin. Chemically or genetically modified mutants areincluded in some embodiments. Various mannanases are known which finduse in the present disclosure (See, e.g., U.S. Pat. Nos. 6,566,114,6,602,842, and 6,440,991, all of which are incorporated herein byreference). Commercially available mannanases that find use in thepresent disclosure include, but are not limited to MANNASTAR®,PURABRITE™, and MANNAWAY®.

Suitable lipases include those of bacterial or fungal origin. Chemicallymodified, proteolytically modified, or protein engineered mutants areincluded. Examples of useful lipases include those from the generaHumicola (e.g., H. lanuginosa, EP258068 and EP305216; H. insolens,WO96/13580), Pseudomonas (e.g., P. alcaligenes or P. pseudoalcaligenes,EP218272; P. cepacia, EP331376; P. stutzeri, GB1372034; P. fluorescensand Pseudomonas sp. strain SD 705, WO95/06720 and WO96/27002; P.wisconsinensis, WO96/12012); and Bacillus (e.g., B. subtilis, Dartois etal., Biochemica et Biophysica Acta 1131:253-360; B. stearothermophilus,JP64/744992; B. pumilus, WO91/16422). Furthermore, a number of clonedlipases find use in some embodiments of the present disclosure,including but not limited to, Penicillium camembertii lipase (See,Yamaguchi et al., Gene 103:61-67 [1991]), Geotricum candidum lipase(See, Schimada et al., J. Biochem., 106:383-388 [1989]), and variousRhizopus lipases such as R. delemar lipase (See, Hass et al., Gene109:117-113 [1991]), a R. niveus lipase (Kugimiya et al., Biosci.Biotech. Biochem. 56:716-719 [1992]) and R. oryzae lipase. Additionallipases useful herein include, for example, those disclosed inWO92/05249, WO94/01541, WO95/35381, WO96/00292, WO95/30744, WO94/25578,WO95/14783, WO95/22615, WO97/04079, WO97/07202, EP407225 and EP260105.Other types of lipase polypeptide enzymes such as cutinases also finduse in some embodiments of the present disclosure, including but notlimited to, cutinase derived from Pseudomonas mendocina (See,WO88/09367), and cutinase derived from Fusarium solani pisi (See,WO90/09446). Examples of certain commercially available lipase enzymesuseful herein include M1 LIPASE™, LUMA FAST™, and LIPOMAX™ (Genencor);LIPEX®, LIPOLASE® and LIPOLASE® ULTRA (Novozymes); and LIPASE P™ “Amano”(Amano Pharmaceutical Co. Ltd., Japan).

Suitable polyesterases include, for example, those disclosed inWO01/34899, WO01/14629 and U.S. Pat. No. 6,933,140.

A detergent composition herein can also comprise 2,6-beta-D-fructanhydrolase, which is effective for removal/cleaning of certain biofilmspresent on household and/or industrial textiles/laundry.

Suitable amylases include, but are not limited to those of bacterial orfungal origin. Chemically or genetically modified mutants are includedin some embodiments. Amylases that find use in the present disclosure,include, but are not limited to, alpha-amylases obtained from B.licheniformis (See e.g., GB1296839). Additional suitable amylasesinclude those disclosed in WO9510603, WO9526397, WO9623874, WO9623873,WO9741213, WO9919467, WO0060060, WO0029560, WO9923211, WO9946399,WO0060058, WO0060059, WO9942567, WO0114532, WO02092797, WO0166712,WO0188107, WO0196537, WO0210355, WO9402597, WO0231124, WO9943793,WO9943794, WO2004113551, WO2005001064, WO2005003311, WO0164852,WO2006063594, WO2006066594, WO2006066596, WO2006012899, WO2008092919,WO2008000825, WO2005018336, WO2005066338, WO2009140504, WO2005019443,WO2010091221, WO2010088447, WO0134784, WO2006012902, WO2006031554,WO2006136161, WO2008101894, WO2010059413, WO2011098531, WO2011080352,WO2011080353, WO2011080354, WO2011082425, WO2011082429, WO2011076123,WO2011087836, WO2011076897, WO94183314, WO9535382, WO9909183, WO9826078,WO9902702, WO9743424, WO9929876, WO9100353, WO9605295, WO9630481,WO9710342, WO2008088493, WO2009149419, WO2009061381, WO2009100102,WO2010104675, WO2010117511, and WO2010115021, all of which areincorporated herein by reference.

Suitable amylases include, for example, commercially available amylasessuch as STAINZYME®, STAINZYME PLUS®, NATALASE®, DURAMYL®, TERMAMYL®,TERMAMYL ULTRA®, FUNGAMYL® and BAN™ (Novo Nordisk A/S and NovozymesA/S); RAPIDASE®, POWERASE®, PURASTAR® and PREFERENZ™ (DuPont IndustrialBiosciences).

Suitable peroxidases/oxidases contemplated for use in the compositionsinclude those of plant, bacterial or fungal origin. Chemically modifiedor protein engineered mutants are included. Examples of peroxidasesuseful herein include those from the genus Coprinus (e.g., C. cinereus,WO93/24618, WO95/10602, and WO98/15257), as well as those referenced inWO2005056782, WO2007106293, WO2008063400, WO2008106214, andWO2008106215. Commercially available peroxidases useful herein include,for example, GUARDZYME™ (Novo Nordisk A/S and Novozymes A/S).

In some embodiments, peroxidases are used in combination with hydrogenperoxide or a source thereof (e.g., a percarbonate, perborate orpersulfate) in the compositions of the present disclosure. In somealternative embodiments, oxidases are used in combination with oxygen.Both types of enzymes are used for “solution bleaching” (i.e., toprevent transfer of a textile dye from a dyed fabric to another fabricwhen the fabrics are washed together in a wash liquor), preferablytogether with an enhancing agent (See e.g., WO94/12621 and WO95/01426).Suitable peroxidases/oxidases include, but are not limited to, those ofplant, bacterial or fungal origin. Chemically or genetically modifiedmutants are included in some embodiments.

Enzymes that may be comprised in a detergent composition herein may bestabilized using conventional stabilizing agents, e.g., a polyol such aspropylene glycol or glycerol; a sugar or sugar alcohol; lactic acid;boric acid or a boric acid derivative (e.g., an aromatic borate ester).

A detergent composition in certain embodiments may comprise one or moreother types of polymers in addition to a dextran as disclosed herein.Examples of other types of polymers useful herein include carboxymethylcellulose (CMC), poly(vinylpyrrolidone) (PVP), polyethylene glycol(PEG), poly(vinyl alcohol) (PVA), polycarboxylates such aspolyacrylates, maleic/acrylic acid copolymers and laurylmethacrylate/acrylic acid copolymers.

A detergent composition herein may contain a bleaching system. Forexample, a bleaching system can comprise an H₂O₂ source such asperborate or percarbonate, which may be combined with a peracid-formingbleach activator such as tetraacetylethylenediamine (TAED) ornonanoyloxybenzenesulfonate (NOBS). Alternatively, a bleaching systemmay comprise peroxyacids (e.g., amide, imide, or sulfone typeperoxyacids). Alternatively still, a bleaching system can be anenzymatic bleaching system comprising perhydrolase, for example, such asthe system described in WO2005/056783.

A detergent composition herein may also contain conventional detergentingredients such as fabric conditioners, clays, foam boosters, sudssuppressors, anti-corrosion agents, soil-suspending agents, anti-soilredeposition agents, dyes, bactericides, tarnish inhibiters, opticalbrighteners, or perfumes. The pH of a detergent composition herein(measured in aqueous solution at use concentration) is usually neutralor alkaline (e.g., pH of about 7.0 to about 11.0).

It is believed that a dextran herein can be included as ananti-redeposition agent and/or clay soil removal agent in a detergentcomposition such as a fabric care composition, if desired (such agentscan optionally be characterized as whiteness maintenance agents incertain aspects). Examples of other suitable anti-redeposition and/orclay soil removal agents herein include polyethoxy zwitterionicsurfactants, water-soluble copolymers of acrylic or methacrylic acidwith acrylic or methacrylic acid-ethylene oxide condensates (e.g., U.S.Pat. No. 3,719,647), cellulose derivatives such ascarboxymethylcellulose and hydroxypropylcellulose (e.g., U.S. Pat. Nos.3,597,416 and 3,523,088), and mixtures comprising nonionic alkylpolyethoxy surfactant, polyethoxy alkyl quaternary cationic surfactantand fatty amide surfactant (e.g., U.S. Pat. No. 4,228,044). Non-limitingexamples of other suitable anti-redeposition and clay soil removalagents are disclosed in U.S. Pat. Nos. 4,597,898 and 4,891,160, and Int.Pat. Appl. Publ. No. WO95/32272, all of which are incorporated herein byreference.

Particular forms of detergent compositions that can be adapted forpurposes disclosed herein are disclosed in, for example,US20090209445A1, US20100081598A1, U.S. Pat. No. 7,001,878B2,EP1504994B1, WO2001085888A2, WO2003089562A1, WO2009098659A1,WO2009098660A1, WO2009112992A1, WO2009124160A1, WO2009152031A1,WO2010059483A1, WO2010088112A1, WO2010090915A1, WO2010135238A1,WO2011094687A1, WO2011094690A1, WO2011127102A1, WO2011163428A1,WO2008000567A1, WO2006045391A1, WO2006007911A1, WO2012027404A1,EP1740690B1, WO2012059336A1, U.S. Pat. No. 6,730,646B1, WO2008087426A1,WO2010116139A1, and WO2012104613A1, all of which are incorporated hereinby reference.

Laundry detergent compositions herein can optionally be heavy duty (allpurpose) laundry detergent compositions. Exemplary heavy duty laundrydetergent compositions comprise a detersive surfactant (10%-40% wt/wt),including an anionic detersive surfactant (selected from a group oflinear or branched or random chain, substituted or unsubstituted alkylsulphates, alkyl sulphonates, alkyl alkoxylated sulphate, alkylphosphates, alkyl phosphonates, alkyl carboxylates, and/or mixturesthereof), and optionally non-ionic surfactant (selected from a group oflinear or branched or random chain, substituted or unsubstituted alkylalkoxylated alcohol, e.g., C8-C18 alkyl ethoxylated alcohols and/orC6-C12 alkyl phenol alkoxylates), where the weight ratio of anionicdetersive surfactant (with a hydrophilic index (Hlc) of from 6.0 to 9)to non-ionic detersive surfactant is greater than 1:1. Suitabledetersive surfactants also include cationic detersive surfactants(selected from a group of alkyl pyridinium compounds, alkyl quaternaryammonium compounds, alkyl quaternary phosphonium compounds, alkylternary sulphonium compounds, and/or mixtures thereof); zwitterionicand/or amphoteric detersive surfactants (selected from a group ofalkanolamine sulpho-betaines); ampholytic surfactants; semi-polarnon-ionic surfactants and mixtures thereof.

A detergent herein such as a heavy duty laundry detergent compositionmay optionally include, a surfactancy boosting polymer consisting ofamphiphilic alkoxylated grease cleaning polymers (selected from a groupof alkoxylated polymers having branched hydrophilic and hydrophobicproperties, such as alkoxylated polyalkylenimines in the range of 0.05wt %-10 wt %) and/or random graft polymers (typically comprising ofhydrophilic backbone comprising monomers selected from the groupconsisting of: unsaturated C1-C6 carboxylic acids, ethers, alcohols,aldehydes, ketones, esters, sugar units, alkoxy units, maleic anhydride,saturated polyalcohols such as glycerol, and mixtures thereof; andhydrophobic side chain(s) selected from the group consisting of: C4-C25alkyl group, polypropylene, polybutylene, vinyl ester of a saturatedC1-C6 mono-carboxylic acid, C1-C6 alkyl ester of acrylic or methacrylicacid, and mixtures thereof.

A detergent herein such as a heavy duty laundry detergent compositionmay optionally include additional polymers such as soil release polymers(include anionically end-capped polyesters, for example SRP1, polymerscomprising at least one monomer unit selected from saccharide,dicarboxylic acid, polyol and combinations thereof, in random or blockconfiguration, ethylene terephthalate-based polymers and co-polymersthereof in random or block configuration, for example REPEL-O-TEX SF,SF-2 AND SRP6, TEXCARE SRA100, SRA300, SRN100, SRN170, SRN240, SRN300AND SRN325, MARLOQUEST SL), anti-redeposition agent(s) herein (0.1 wt %to 10 wt %), include carboxylate polymers, such as polymers comprisingat least one monomer selected from acrylic acid, maleic acid (or maleicanhydride), fumaric acid, itaconic acid, aconitic acid, mesaconic acid,citraconic acid, methylenemalonic acid, and any mixture thereof,vinylpyrrolidone homopolymer, and/or polyethylene glycol, molecularweight in the range of from 500 to 100,000 Da); and polymericcarboxylate (such as maleate/acrylate random copolymer or polyacrylatehomopolymer).

A detergent herein such as a heavy duty laundry detergent compositionmay optionally further include saturated or unsaturated fatty acids,preferably saturated or unsaturated C12-C24 fatty acids (0 wt % to 10 wt%); deposition aids in addition to a dextran compound disclosed herein(examples for which include polysaccharides, cellulosic polymers, polydiallyl dimethyl ammonium halides (DADMAC), and co-polymers of DAD MACwith vinyl pyrrolidone, acrylamides, imidazoles, imidazolinium halides,and mixtures thereof, in random or block configuration, cationic guargum, cationic starch, cationic polyacylamides, and mixtures thereof.

A detergent herein such as a heavy duty laundry detergent compositionmay optionally further include dye transfer inhibiting agents, examplesof which include manganese phthalocyanine, peroxidases,polyvinylpyrrolidone polymers, polyamine N-oxide polymers, copolymers ofN-vinylpyrrolidone and N-vinylimidazole, polyvinyloxazolidones andpolyvinylimidazoles and/or mixtures thereof; chelating agents, examplesof which include ethylene-diamine-tetraacetic acid (EDTA), diethylenetriamine penta methylene phosphonic acid (DTPMP), hydroxy-ethanediphosphonic acid (HEDP), ethylenediamine N,N′-disuccinic acid (EDDS),methyl glycine diacetic acid (MGDA), diethylene triamine penta aceticacid (DTPA), propylene diamine tetracetic acid (PDTA),2-hydroxypyridine-N-oxide (HPNO), or methyl glycine diacetic acid(MGDA), glutamic acid N,N-diacetic acid (N,N-dicarboxymethyl glutamicacid tetrasodium salt (GLDA), nitrilotriacetic acid (NTA),4,5-dihydroxy-m-benzenedisulfonic acid, citric acid and any saltsthereof, N-hydroxyethylethylenediaminetriacetic acid (HEDTA),triethylenetetraaminehexaacetic acid (TTNA), N-hydroxyethyliminodiaceticacid (HEIDA), dihydroxyethylglycine (DHEG),ethylenediaminetetrapropionic acid (EDTP), and derivatives thereof.

A detergent herein such as a heavy duty laundry detergent compositionmay optionally include silicone or fatty-acid based suds suppressors;hueing dyes, calcium and magnesium cations, visual signalingingredients, anti-foam (0.001 wt % to about 4.0 wt %), and/or astructurant/thickener (0.01 wt % to 5 wt %) selected from the groupconsisting of diglycerides and triglycerides, ethylene glycoldistearate, microcrystalline cellulose, microfiber cellulose,biopolymers, xanthan gum, gellan gum, and mixtures thereof). Suchstructurant/thickener would be, in certain embodiments, in addition tothe one or more dextran compounds comprised in the detergent. Astructurant can also be referred to as a structural agent.

A detergent herein can be in the form of a heavy duty dry/solid laundrydetergent composition, for example. Such a detergent may include: (i) adetersive surfactant, such as any anionic detersive surfactant disclosedherein, any non-ionic detersive surfactant disclosed herein, anycationic detersive surfactant disclosed herein, any zwitterionic and/oramphoteric detersive surfactant disclosed herein, any ampholyticsurfactant, any semi-polar non-ionic surfactant, and mixtures thereof;(ii) a builder, such as any phosphate-free builder (e.g., zeolitebuilders in the range of 0 wt % to less than 10 wt %), any phosphatebuilder (e.g., sodium tri-polyphosphate in the range of 0 wt % to lessthan 10 wt %), citric acid, citrate salts and nitrilotriacetic acid, anysilicate salt (e.g., sodium or potassium silicate or sodiummeta-silicate in the range of 0 wt % to less than 10 wt %); anycarbonate salt (e.g., sodium carbonate and/or sodium bicarbonate in therange of 0 wt % to less than 80 wt %), and mixtures thereof; (iii) ableaching agent, such as any photobleach (e.g., sulfonated zincphthalocyanines, sulfonated aluminum phthalocyanines, xanthenes dyes,and mixtures thereof), any hydrophobic or hydrophilic bleach activator(e.g., dodecanoyl oxybenzene sulfonate, decanoyl oxybenzene sulfonate,decanoyl oxybenzoic acid or salts thereof, 3,5,5-trimethy hexanoyloxybenzene sulfonate, tetraacetyl ethylene diamine-TAED,nonanoyloxybenzene sulfonate-NOBS, nitrile quats, and mixtures thereof),any source of hydrogen peroxide (e.g., inorganic perhydrate salts,examples of which include mono or tetra hydrate sodium salt ofperborate, percarbonate, persulfate, perphosphate, or persilicate), anypreformed hydrophilic and/or hydrophobic peracids (e.g., percarboxylicacids and salts, percarbonic acids and salts, perimidic acids and salts,peroxymonosulfuric acids and salts, and mixtures thereof); and/or (iv)any other components such as a bleach catalyst (e.g., imine bleachboosters examples of which include iminium cations and polyions, iminiumzwitterions, modified amines, modified amine oxides, N-sulphonyl imines,N-phosphonyl imines, N-acyl imines, thiadiazole dioxides,perfluoroimines, cyclic sugar ketones, and mixtures thereof), and ametal-containing bleach catalyst (e.g., copper, iron, titanium,ruthenium, tungsten, molybdenum, or manganese cations along with anauxiliary metal cations such as zinc or aluminum and a sequestrate suchas EDTA, ethylenediaminetetra (methylenephosphonic acid).

Compositions disclosed herein can be in the form of a dishwashingdetergent composition, for example. Examples of dishwashing detergentsinclude automatic dishwashing detergents (typically used in dishwashermachines) and hand-washing dish detergents. A dishwashing detergentcomposition can be in any dry or liquid/aqueous form as disclosedherein, for example. Components that may be included in certainembodiments of a dishwashing detergent composition include, for example,one or more of a phosphate; oxygen- or chlorine-based bleaching agent;non-ionic surfactant; alkaline salt (e.g., metasilicates, alkali metalhydroxides, sodium carbonate); any active enzyme disclosed herein;anti-corrosion agent (e.g., sodium silicate); anti-foaming agent;additives to slow down the removal of glaze and patterns from ceramics;perfume; anti-caking agent (in granular detergent); starch (intablet-based detergents); gelling agent (in liquid/gel baseddetergents); and/or sand (powdered detergents).

Dishwashing detergents such as an automatic dishwasher detergent orliquid dishwashing detergent can comprise (i) a non-ionic surfactant,including any ethoxylated non-ionic surfactant, alcohol alkoxylatedsurfactant, epoxy-capped poly(oxyalkylated) alcohol, or amine oxidesurfactant present in an amount from 0 to 10 wt %; (ii) a builder, inthe range of about 5-60 wt %, including any phosphate builder (e.g.,mono-phosphates, di-phosphates, tri-polyphosphates, otheroligomeric-polyphosphates, sodium tripolyphosphate-STPP), anyphosphate-free builder (e.g., amino acid-based compounds includingmethyl-glycine-diacetic acid [MGDA] and salts or derivatives thereof,glutamic-N,N-diacetic acid [GLDA] and salts or derivatives thereof,iminodisuccinic acid (IDS) and salts or derivatives thereof, carboxymethyl inulin and salts or derivatives thereof, nitrilotriacetic acid[NTA], diethylene triamine penta acetic acid [DTPA], B-alaninediaceticacid [B-ADA] and salts thereof), homopolymers and copolymers ofpoly-carboxylic acids and partially or completely neutralized saltsthereof, monomeric polycarboxylic acids and hydroxycarboxylic acids andsalts thereof in the range of 0.5 wt % to 50 wt %, orsulfonated/carboxylated polymers in the range of about 0.1 wt % to about50 wt %; (iii) a drying aid in the range of about 0.1 wt % to about 10wt % (e.g., polyesters, especially anionic polyesters, optionallytogether with further monomers with 3 to 6 functionalities—typicallyacid, alcohol or ester functionalities which are conducive topolycondensation, polycarbonate-, polyurethane- and/orpolyurea-polyorganosiloxane compounds or precursor compounds thereof,particularly of the reactive cyclic carbonate and urea type); (iv) asilicate in the range from about 1 wt % to about 20 wt % (e.g., sodiumor potassium silicates such as sodium disilicate, sodium meta-silicateand crystalline phyllosilicates); (v) an inorganic bleach (e.g.,perhydrate salts such as perborate, percarbonate, perphosphate,persulfate and persilicate salts) and/or an organic bleach (e.g.,organic peroxyacids such as diacyl- and tetraacylperoxides, especiallydiperoxydodecanedioic acid, diperoxytetradecanedioic acid, anddiperoxyhexadecanedioic acid); (vi) a bleach activator (e.g., organicperacid precursors in the range from about 0.1 wt % to about 10 wt %)and/or bleach catalyst (e.g., manganese triazacyclononane and relatedcomplexes; Co, Cu, Mn, and Fe bispyridylamine and related complexes; andpentamine acetate cobalt (III) and related complexes); (vii) a metalcare agent in the range from about 0.1 wt % to 5 wt % (e.g.,benzatriazoles, metal salts and complexes, and/or silicates); and/or(viii) any active enzyme disclosed herein in the range from about 0.01to 5.0 mg of active enzyme per gram of automatic dishwashing detergentcomposition, and an enzyme stabilizer component (e.g., oligosaccharides,polysaccharides, and inorganic divalent metal salts).

Various examples of detergent formulations comprising at least onedextran herein are disclosed below (1-19):

1) A detergent composition formulated as a granulate having a bulkdensity of at least 600 g/L comprising: linear alkylbenzenesulfonate(calculated as acid) at about 7-12 wt %; alcohol ethoxysulfate (e.g.,C12-18 alcohol, 1-2 ethylene oxide [EO]) or alkyl sulfate (e.g., C16-18)at about 1-4 wt %; alcohol ethoxylate (e.g., C14-15 alcohol) at about5-9 wt %; sodium carbonate at about 14-20 wt %; soluble silicate (e.g.,Na₂O 2SiO₂) at about 2-6 wt %; zeolite (e.g., NaAlSiO₄) at about 15-22wt %; sodium sulfate at about 0-6 wt %; sodium citrate/citric acid atabout 0-15 wt %; sodium perborate at about 11-18 wt %; TAED at about 2-6wt %; dextran herein up to about 2 wt %; other polymers (e.g.,maleic/acrylic acid copolymer, PVP, PEG) at about 0-3 wt %; optionallyan enzyme(s) (calculated as pure enzyme protein) at about 0.0001-0.1 wt%; and minor ingredients (e.g., suds suppressors, perfumes, opticalbrightener, photobleach) at about 0-5 wt %.

2) A detergent composition formulated as a granulate having a bulkdensity of at least 600 g/L comprising: linear alkylbenzenesulfonate(calculated as acid) at about 6-11 wt %; alcohol ethoxysulfate (e.g.,C12-18 alcohol, 1-2 EO) or alkyl sulfate (e.g., C16-18) at about 1-3 wt%; alcohol ethoxylate (e.g., C14-15 alcohol) at about 5-9 wt %; sodiumcarbonate at about 15-21 wt %; soluble silicate (e.g., Na₂O 2SiO₂) atabout 1-4 wt %; zeolite (e.g., NaAlSiO₄) at about 24-34 wt %; sodiumsulfate at about 4-10 wt %; sodium citrate/citric acid at about 0-15 wt%; sodium perborate at about 11-18 wt %; TAED at about 2-6 wt %; dextranherein up to about 2 wt %; other polymers (e.g., maleic/acrylic acidcopolymer, PVP, PEG) at about 1-6 wt %; optionally an enzyme(s)(calculated as pure enzyme protein) at about 0.0001-0.1 wt %; and minoringredients (e.g., suds suppressors, perfumes, optical brightener,photobleach) at about 0-5 wt %.

3) A detergent composition formulated as a granulate having a bulkdensity of at least 600 g/L comprising: linear alkylbenzenesulfonate(calculated as acid) at about 5-9 wt %; alcohol ethoxysulfate (e.g.,C12-18 alcohol, 7 EO) at about 7-14 wt %; soap as fatty acid (e.g.,C16-22 fatty acid) at about 1-3 wt %; sodium carbonate at about 10-17 wt%; soluble silicate (e.g., Na₂O 2SiO₂) at about 3-9 wt %; zeolite (e.g.,NaAlSiO₄) at about 23-33 wt %; sodium sulfate at about 0-4 wt %; sodiumperborate at about 8-16 wt %; TAED at about 2-8 wt %; phosphonate (e.g.,EDTMPA) at about 0-1 wt %; dextran herein up to about 2 wt %; otherpolymers (e.g., maleic/acrylic acid copolymer, PVP, PEG) at about 0-3 wt%; optionally an enzyme(s) (calculated as pure enzyme protein) at about0.0001-0.1 wt %; and minor ingredients (e.g., suds suppressors,perfumes, optical brightener) at about 0-5 wt %.

4) A detergent composition formulated as a granulate having a bulkdensity of at least 600 g/L comprising: linear alkylbenzenesulfonate(calculated as acid) at about 8-12 wt %; alcohol ethoxylate (e.g.,C12-18 alcohol, 7 EO) at about 10-25 wt %; sodium carbonate at about14-22 wt %; soluble silicate (e.g., Na₂O 2SiO₂) at about 1-5 wt %;zeolite (e.g., NaAlSiO₄) at about 25-35 wt %; sodium sulfate at about0-10 wt %; sodium perborate at about 8-16 wt %; TAED at about 2-8 wt %;phosphonate (e.g., EDTMPA) at about 0-1 wt %; dextran herein up to about2 wt %; other polymers (e.g., maleic/acrylic acid copolymer, PVP, PEG)at about 1-3 wt %; optionally an enzyme(s) (calculated as pure enzymeprotein) at about 0.0001-0.1 wt %; and minor ingredients (e.g., sudssuppressors, perfumes) at about 0-5 wt %.

5) An aqueous liquid detergent composition comprising: linearalkylbenzenesulfonate (calculated as acid) at about 15-21 wt %; alcoholethoxylate (e.g., C12-18 alcohol, 7 EO; or C12-15 alcohol, 5 EO) atabout 12-18 wt %; soap as fatty acid (e.g., oleic acid) at about 3-13 wt%; alkenylsuccinic acid (C12-14) at about 0-13 wt %; aminoethanol atabout 8-18 wt %; citric acid at about 2-8 wt %; phosphonate at about 0-3wt %; dextran herein up to about 2 wt %; other polymers (e.g., PVP, PEG)at about 0-3 wt %; borate at about 0-2 wt %; ethanol at about 0-3 wt %;propylene glycol at about 8-14 wt %; optionally an enzyme(s) (calculatedas pure enzyme protein) at about 0.0001-0.1 wt %; and minor ingredients(e.g., dispersants, suds suppressors, perfume, optical brightener) atabout 0-5 wt %.

6) An aqueous structured liquid detergent composition comprising: linearalkylbenzenesulfonate (calculated as acid) at about 15-21 wt %; alcoholethoxylate (e.g., C12-18 alcohol, 7 EO; or C12-15 alcohol, 5 EO) atabout 3-9 wt %; soap as fatty acid (e.g., oleic acid) at about 3-10 wt%; zeolite (e.g., NaAlSiO₄) at about 14-22 wt %; potassium citrate about9-18 wt %; borate at about 0-2 wt %; dextran herein up to about 2 wt %;other polymers (e.g., PVP, PEG) at about 0-3 wt %; ethanol at about 0-3wt %; anchoring polymers (e.g., lauryl methacrylate/acrylic acidcopolymer, molar ratio 25:1, MW 3800) at about 0-3 wt %; glycerol atabout 0-5 wt %; optionally an enzyme(s) (calculated as pure enzymeprotein) at about 0.0001-0.1 wt %; and minor ingredients (e.g.,dispersants, suds suppressors, perfume, optical brightener) at about 0-5wt %.

7) A detergent composition formulated as a granulate having a bulkdensity of at least 600 g/L comprising: fatty alcohol sulfate at about5-10 wt %, ethoxylated fatty acid monoethanolamide at about 3-9 wt %;soap as fatty acid at about 0-3 wt %; sodium carbonate at about 5-10 wt%; soluble silicate (e.g., Na₂O 2SiO₂) at about 1-4 wt %; zeolite (e.g.,NaAlSiO₄) at about 20-40 wt %; sodium sulfate at about 2-8 wt %; sodiumperborate at about 12-18 wt %; TAED at about 2-7 wt %; dextran herein upto about 2 wt %; other polymers (e.g., maleic/acrylic acid copolymer,PEG) at about 1-5 wt %; optionally an enzyme(s) (calculated as pureenzyme protein) at about 0.0001-0.1 wt %; and minor ingredients (e.g.,optical brightener, suds suppressors, perfumes) at about 0-5 wt %.

8) A detergent composition formulated as a granulate comprising: linearalkylbenzenesulfonate (calculated as acid) at about 8-14 wt %;ethoxylated fatty acid monoethanolamide at about 5-11 wt %; soap asfatty acid at about 0-3 wt %; sodium carbonate at about 4-10 wt %;soluble silicate (e.g., Na₂O 2SiO₂) at about 1-4 wt %; zeolite (e.g.,NaAlSiO₄) at about 30-50 wt %; sodium sulfate at about 3-11 wt %; sodiumcitrate at about 5-12 wt %; dextran herein up to about 2 wt %; otherpolymers (e.g., PVP, maleic/acrylic acid copolymer, PEG) at about 1-5 wt%; optionally an enzyme(s) (calculated as pure enzyme protein) at about0.0001-0.1 wt %; and minor ingredients (e.g., suds suppressors,perfumes) at about 0-5 wt %.

9) A detergent composition formulated as a granulate comprising: linearalkylbenzenesulfonate (calculated as acid) at about 6-12 wt %; nonionicsurfactant at about 1-4 wt %; soap as fatty acid at about 2-6 wt %;sodium carbonate at about 14-22 wt %; zeolite (e.g., NaAlSiO₄) at about18-32 wt %; sodium sulfate at about 5-20 wt %; sodium citrate at about3-8 wt %; sodium perborate at about 4-9 wt %; bleach activator (e.g.,NOBS or TAED) at about 1-5 wt %; dextran herein up to about 2 wt %;other polymers (e.g., polycarboxylate or PEG) at about 1-5 wt %;optionally an enzyme(s) (calculated as pure enzyme protein) at about0.0001-0.1 wt %; and minor ingredients (e.g., optical brightener,perfume) at about 0-5 wt %.

10) An aqueous liquid detergent composition comprising: linearalkylbenzenesulfonate (calculated as acid) at about 15-23 wt %; alcoholethoxysulfate (e.g., C12-15 alcohol, 2-3 EO) at about 8-15 wt %; alcoholethoxylate (e.g., C12-15 alcohol, 7 EO; or C12-15 alcohol, 5 EO) atabout 3-9 wt %; soap as fatty acid (e.g., lauric acid) at about 0-3 wt%; aminoethanol at about 1-5 wt %; sodium citrate at about 5-10 wt %;hydrotrope (e.g., sodium toluenesulfonate) at about 2-6 wt %; borate atabout 0-2 wt %; dextran herein up to about 1 wt %; ethanol at about 1-3wt %; propylene glycol at about 2-5 wt %; optionally an enzyme(s)(calculated as pure enzyme protein) at about 0.0001-0.1 wt %; and minoringredients (e.g., dispersants, perfume, optical brighteners) at about0-5 wt %.

11) An aqueous liquid detergent composition comprising: linearalkylbenzenesulfonate (calculated as acid) at about 20-32 wt %; alcoholethoxylate (e.g., C12-15 alcohol, 7 EO; or C12-15 alcohol, 5 EO) atabout 6-12 wt %; aminoethanol at about 2-6 wt %; citric acid at about8-14 wt %; borate at about 1-3 wt %; dextran herein up to about 2 wt %;ethanol at about 1-3 wt %; propylene glycol at about 2-5 wt %; otherpolymers (e.g., maleic/acrylic acid copolymer, anchoring polymer such aslauryl methacrylate/acrylic acid copolymer) at about 0-3 wt %; glycerolat about 3-8 wt %; optionally an enzyme(s) (calculated as pure enzymeprotein) at about 0.0001-0.1 wt %; and minor ingredients (e.g.,hydrotropes, dispersants, perfume, optical brighteners) at about 0-5 wt%.

12) A detergent composition formulated as a granulate having a bulkdensity of at least 600 g/L comprising: anionic surfactant (e.g., linearalkylbenzenesulfonate, alkyl sulfate, alpha-olefinsulfonate, alpha-sulfofatty acid methyl esters, alkanesulfonates, soap) at about 25-40 wt %;nonionic surfactant (e.g., alcohol ethoxylate) at about 1-10 wt %;sodium carbonate at about 8-25 wt %; soluble silicate (e.g., Na₂O 2SiO₂)at about 5-15 wt %; sodium sulfate at about 0-5 wt %; zeolite (NaAlSiO₄)at about 15-28 wt %; sodium perborate at about 0-20 wt %; bleachactivator (e.g., TAED or NOBS) at about 0-5 wt %; dextran herein up toabout 2 wt %; optionally an enzyme(s) (calculated as pure enzymeprotein) at about 0.0001-0.1 wt %; and minor ingredients (e.g., perfume,optical brighteners) at about 0-3 wt %.

13) Detergent compositions as described in (1)-(12) above, but in whichall or part of the linear alkylbenzenesulfonate is replaced by C12-C18alkyl sulfate.

14) A detergent composition formulated as a granulate having a bulkdensity of at least 600 g/L comprising: C12-C18 alkyl sulfate at about9-15 wt %; alcohol ethoxylate at about 3-6 wt %; polyhydroxy alkyl fattyacid amide at about 1-5 wt %; zeolite (e.g., NaAlSiO₄) at about 10-20 wt%; layered disilicate (e.g., SK56 from Hoechst) at about 10-20 wt %;sodium carbonate at about 3-12 wt %; soluble silicate (e.g., Na₂O 2SiO₂)at 0-6 wt %; sodium citrate at about 4-8 wt %; sodium percarbonate atabout 13-22 wt %; TAED at about 3-8 wt %; dextran herein up to about 2wt %; other polymers (e.g., polycarboxylates and PVP) at about 0-5 wt %;optionally an enzyme(s) (calculated as pure enzyme protein) at about0.0001-0.1 wt %; and minor ingredients (e.g., optical brightener,photobleach, perfume, suds suppressors) at about 0-5 wt %.

15) A detergent composition formulated as a granulate having a bulkdensity of at least 600 g/L comprising: C12-C18 alkyl sulfate at about4-8 wt %; alcohol ethoxylate at about 11-15 wt %; soap at about 1-4 wt%; zeolite MAP or zeolite A at about 35-45 wt %; sodium carbonate atabout 2-8 wt %; soluble silicate (e.g., Na₂O 2SiO₂) at 0-4 wt %; sodiumpercarbonate at about 13-22 wt %; TAED at about 1-8 wt %; dextran hereinup to about 3 wt %; other polymers (e.g., polycarboxylates and PVP) atabout 0-3 wt %; optionally an enzyme(s) (calculated as pure enzymeprotein) at about 0.0001-0.1 wt %; and minor ingredients (e.g., opticalbrightener, phosphonate, perfume) at about 0-3 wt %.

16) Detergent formulations as described in (1)-(15) above, but thatcontain a stabilized or encapsulated peracid, either as an additionalcomponent or as a substitute for an already specified bleach system(s).

17) Detergent compositions as described in (1), (3), (7), (9) and (12)above, but in which perborate is replaced by percarbonate.

18) Detergent compositions as described in (1), (3), (7), (9), (12),(14) and (15) above, but that additionally contain a manganese catalyst.A manganese catalyst, for example, is one of the compounds described byHage et al. (1994, Nature 369:637-639), which is incorporated herein byreference.

19) Detergent compositions formulated as a non-aqueous detergent liquidcomprising a liquid non-ionic surfactant (e.g., a linear alkoxylatedprimary alcohol), a builder system (e.g., phosphate), dextran herein,optionally an enzyme(s), and alkali. The detergent may also comprise ananionic surfactant and/or bleach system.

It is believed that numerous commercially available detergentformulations can be adapted to include a dextran compound disclosedherein. Examples include PUREX® ULTRAPACKS (Henkel), FINISH® QUANTUM(Reckitt Benckiser), CLOROX™ 2 PACKS (Clorox), OXICLEAN MAX FORCE POWERPAKS (Church & Dwight), TIDE® STAIN RELEASE, CASCADE® ACTIONPACS, andTIDE® PODS™ (Procter & Gamble).

Compositions disclosed herein can be in the form of an oral carecomposition, for example. Examples of oral care compositions includedentifrices, toothpaste, mouth wash, mouth rinse, chewing gum, andedible strips that provide some form of oral care (e.g., treatment orprevention of cavities [dental caries], gingivitis, plaque, tartar,and/or periodontal disease). An oral care composition can also be fortreating an “oral surface”, which encompasses any soft or hard surfacewithin the oral cavity including surfaces of the tongue, hard and softpalate, buccal mucosa, gums and dental surfaces. A “dental surface”herein is a surface of a natural tooth or a hard surface of artificialdentition including a crown, cap, filling, bridge, denture, or dentalimplant, for example.

An oral care composition herein can comprise about 0.01-15.0 wt % (e.g.,˜0.1-10 wt % or ˜0.1-5.0 wt %, ˜0.1-2.0 wt %) of one or more dextranether compounds as disclosed herein, for example. One or more dextranether compounds comprised in an oral care composition can sometimes beprovided therein as a thickening agent and/or dispersion agent, whichmay be useful to impart a desired consistency and/or mouth feel to thecomposition. One or more other thickening or dispersion agents can alsobe provided in an oral care composition herein, such as a carboxyvinylpolymer, carrageenan (e.g., L-carrageenan), natural gum (e.g., karaya,xanthan, gum arabic, tragacanth), colloidal magnesium aluminum silicate,or colloidal silica, for example.

An oral care composition herein may be a toothpaste or other dentifrice,for example. Such compositions, as well as any other oral carecomposition herein, can additionally comprise, without limitation, oneor more of an anticaries agent, antimicrobial or antibacterial agent,anticalculus or tartar control agent, surfactant, abrasive, pH-modifyingagent, foam modulator, humectant, flavorant, sweetener,pigment/colorant, whitening agent, and/or other suitable components.Examples of oral care compositions to which one or more dextrancompounds can be added are disclosed in U.S. Patent Appl. Publ. Nos.2006/0134025, 2002/0022006 and 2008/0057007, which are incorporatedherein by reference.

An anticaries agent herein can be an orally acceptable source offluoride ions. Suitable sources of fluoride ions include fluoride,monofluorophosphate and fluorosilicate salts as well as amine fluorides,including olaflur(N′-octadecyltrimethylendiamine-N,N,N′-tris(2-ethanol)-dihydrofluoride),for example. An anticaries agent can be present in an amount providing atotal of about 100-20000 ppm, about 200-5000 ppm, or about 500-2500 ppm,fluoride ions to the composition, for example. In oral care compositionsin which sodium fluoride is the sole source of fluoride ions, an amountof about 0.01-5.0 wt %, about 0.05-1.0 wt %, or about 0.1-0.5 wt %,sodium fluoride can be present in the composition, for example.

An antimicrobial or antibacterial agent suitable for use in an oral carecomposition herein includes, for example, phenolic compounds (e.g.,4-allylcatechol; p-hydroxybenzoic acid esters such as benzylparaben,butylparaben, ethylparaben, methylparaben and propylparaben;2-benzylphenol; butylated hydroxyanisole; butylated hydroxytoluene;capsaicin; carvacrol; creosol; eugenol; guaiacol; halogenatedbisphenolics such as hexachlorophene and bromochlorophene;4-hexylresorcinol; 8-hydroxyquinoline and salts thereof; salicylic acidesters such as menthyl salicylate, methyl salicylate and phenylsalicylate; phenol; pyrocatechol; salicylanilide; thymol; halogenateddiphenylether compounds such as triclosan and triclosan monophosphate),copper (II) compounds (e.g., copper (II) chloride, fluoride, sulfate andhydroxide), zinc ion sources (e.g., zinc acetate, citrate, gluconate,glycinate, oxide, and sulfate), phthalic acid and salts thereof (e.g.,magnesium monopotassium phthalate), hexetidine, octenidine,sanguinarine, benzalkonium chloride, domiphen bromide, alkylpyridiniumchlorides (e.g. cetylpyridinium chloride, tetradecylpyridinium chloride,N-tetradecyl-4-ethylpyridinium chloride), iodine, sulfonamides,bisbiguanides (e.g., alexidine, chlorhexidine, chlorhexidinedigluconate), piperidino derivatives (e.g., delmopinol, octapinol),magnolia extract, grapeseed extract, rosemary extract, menthol,geraniol, citral, eucalyptol, antibiotics (e.g., augmentin, amoxicillin,tetracycline, doxycycline, minocycline, metronidazole, neomycin,kanamycin, clindamycin), and/or any antibacterial agents disclosed inU.S. Pat. No. 5,776,435, which is incorporated herein by reference. Oneor more antimicrobial agents can optionally be present at about 0.01-10wt % (e.g., 0.1-3 wt %), for example, in the disclosed oral carecomposition.

An anticalculus or tartar control agent suitable for use in an oral carecomposition herein includes, for example, phosphates and polyphosphates(e.g., pyrophosphates), polyaminopropanesulfonic acid (AMPS), zinccitrate trihydrate, polypeptides (e.g., polyaspartic and polyglutamicacids), polyolefin sulfonates, polyolefin phosphates, diphosphonates(e.g., azacycloalkane-2,2-diphosphonates such asazacycloheptane-2,2-diphosphonic acid), N-methylazacyclopentane-2,3-diphosphonic acid, ethane-1-hydroxy-1,1-diphosphonicacid (EHDP), ethane-1-amino-1,1-diphosphonate, and/or phosphonoalkanecarboxylic acids and salts thereof (e.g., their alkali metal andammonium salts). Useful inorganic phosphate and polyphosphate saltsinclude, for example, monobasic, dibasic and tribasic sodium phosphates,sodium tripolyphosphate, tetrapolyphosphate, mono-, di-, tri- andtetra-sodium pyrophosphates, disodium dihydrogen pyrophosphate, sodiumtrimetaphosphate, sodium hexametaphosphate, or any of these in whichsodium is replaced by potassium or ammonium. Other useful anticalculusagents in certain embodiments include anionic polycarboxylate polymers(e.g., polymers or copolymers of acrylic acid, methacrylic, and maleicanhydride such as polyvinyl methyl ether/maleic anhydride copolymers).Still other useful anticalculus agents include sequestering agents suchas hydroxycarboxylic acids (e.g., citric, fumaric, malic, glutaric andoxalic acids and salts thereof) and aminopolycarboxylic acids (e.g.,EDTA). One or more anticalculus or tartar control agents can optionallybe present at about 0.01-50 wt % (e.g., about 0.05-25 wt % or about0.1-15 wt %), for example, in the disclosed oral care composition.

A surfactant suitable for use in an oral care composition herein may beanionic, non-ionic, or amphoteric, for example. Suitable anionicsurfactants include, without limitation, water-soluble salts of C₈₋₂₀alkyl sulfates, sulfonated monoglycerides of C₈₋₂₀ fatty acids,sarcosinates, and taurates. Examples of anionic surfactants includesodium lauryl sulfate, sodium coconut monoglyceride sulfonate, sodiumlauryl sarcosinate, sodium lauryl isoethionate, sodium laurethcarboxylate and sodium dodecyl benzenesulfonate. Suitable non-ionicsurfactants include, without limitation, poloxamers, polyoxyethylenesorbitan esters, fatty alcohol ethoxylates, alkylphenol ethoxylates,tertiary amine oxides, tertiary phosphine oxides, and dialkylsulfoxides. Suitable amphoteric surfactants include, without limitation,derivatives of C₈₋₂₀ aliphatic secondary and tertiary amines having ananionic group such as a carboxylate, sulfate, sulfonate, phosphate orphosphonate. An example of a suitable amphoteric surfactant iscocoamidopropyl betaine. One or more surfactants are optionally presentin a total amount of about 0.01-10 wt % (e.g., about 0.05-5.0 wt % orabout 0.1-2.0 wt %), for example, in the disclosed oral carecomposition.

An abrasive suitable for use in an oral care composition herein mayinclude, for example, silica (e.g., silica gel, hydrated silica,precipitated silica), alumina, insoluble phosphates, calcium carbonate,and resinous abrasives (e.g., a urea-formaldehyde condensation product).Examples of insoluble phosphates useful as abrasives herein areorthophosphates, polymetaphosphates and pyrophosphates, and includedicalcium orthophosphate dihydrate, calcium pyrophosphate, beta-calciumpyrophosphate, tricalcium phosphate, calcium polymetaphosphate andinsoluble sodium polymetaphosphate. One or more abrasives are optionallypresent in a total amount of about 5-70 wt % (e.g., about 10-56 wt % orabout 15-30 wt %), for example, in the disclosed oral care composition.The average particle size of an abrasive in certain embodiments is about0.1-30 microns (e.g., about 1-20 microns or about 5-15 microns).

An oral care composition in certain embodiments may comprise at leastone pH-modifying agent. Such agents may be selected to acidify, makemore basic, or buffer the pH of a composition to a pH range of about2-10 (e.g., pH ranging from about 2-8, 3-9, 4-8, 5-7, 6-10, or 7-9).Examples of pH-modifying agents useful herein include, withoutlimitation, carboxylic, phosphoric and sulfonic acids; acid salts (e.g.,monosodium citrate, disodium citrate, monosodium malate); alkali metalhydroxides (e.g. sodium hydroxide, carbonates such as sodium carbonate,bicarbonates, sesquicarbonates); borates; silicates; phosphates (e.g.,monosodium phosphate, trisodium phosphate, pyrophosphate salts); andimidazole.

A foam modulator suitable for use in an oral care composition herein maybe a polyethylene glycol (PEG), for example. High molecular weight PEGsare suitable, including those having an average molecular weight ofabout 200000-7000000 (e.g., about 500000-5000000 or about1000000-2500000), for example. One or more PEGs are optionally presentin a total amount of about 0.1-10 wt % (e.g. about 0.2-5.0 wt % or about0.25-2.0 wt %), for example, in the disclosed oral care composition.

An oral care composition in certain embodiments may comprise at leastone humectant. A humectant in certain embodiments may be a polyhydricalcohol such as glycerin, sorbitol, xylitol, or a low molecular weightPEG. Most suitable humectants also may function as a sweetener herein.One or more humectants are optionally present in a total amount of about1.0-70 wt % (e.g., about 1.0-50 wt %, about 2-25 wt %, or about 5-15 wt%), for example, in the disclosed oral care composition.

A natural or artificial sweetener may optionally be comprised in an oralcare composition herein. Examples of suitable sweeteners includedextrose, sucrose, maltose, dextrin, invert sugar, mannose, xylose,ribose, fructose, levulose, galactose, corn syrup (e.g., high fructosecorn syrup or corn syrup solids), partially hydrolyzed starch,hydrogenated starch hydrolysate, sorbitol, mannitol, xylitol, maltitol,isomalt, aspartame, neotame, saccharin and salts thereof,dipeptide-based intense sweeteners, and cyclamates. One or moresweeteners are optionally present in a total amount of about 0.005-5.0wt %, for example, in the disclosed oral care composition.

A natural or artificial flavorant may optionally be comprised in an oralcare composition herein. Examples of suitable flavorants includevanillin; sage; marjoram; parsley oil; spearmint oil; cinnamon oil; oilof wintergreen (methylsalicylate); peppermint oil; clove oil; bay oil;anise oil; eucalyptus oil; citrus oils; fruit oils; essences such asthose derived from lemon, orange, lime, grapefruit, apricot, banana,grape, apple, strawberry, cherry, or pineapple; bean- and nut-derivedflavors such as coffee, cocoa, cola, peanut, or almond; and adsorbed andencapsulated flavorants. Also encompassed within flavorants herein areingredients that provide fragrance and/or other sensory effect in themouth, including cooling or warming effects. Such ingredients include,without limitation, menthol, menthyl acetate, menthyl lactate, camphor,eucalyptus oil, eucalyptol, anethole, eugenol, cassia, oxanone,Irisone®, propenyl guaiethol, thymol, linalool, benzaldehyde,cinnamaldehyde, N-ethyl-p-menthan-3-carboxamine,N,2,3-trimethyl-2-isopropylbutanamide, 3-(1-menthoxy)-propane-1,2-diol,cinnamaldehyde glycerol acetal (CGA), and menthone glycerol acetal(MGA). One or more flavorants are optionally present in a total amountof about 0.01-5.0 wt % (e.g., about 0.1-2.5 wt %), for example, in thedisclosed oral care composition.

An oral care composition in certain embodiments may comprise at leastone bicarbonate salt. Any orally acceptable bicarbonate can be used,including alkali metal bicarbonates such as sodium or potassiumbicarbonate, and ammonium bicarbonate, for example. One or morebicarbonate salts are optionally present in a total amount of about0.1-50 wt % (e.g., about 1-20 wt %), for example, in the disclosed oralcare composition.

An oral care composition in certain embodiments may comprise at leastone whitening agent and/or colorant. A suitable whitening agent is aperoxide compound such as any of those disclosed in U.S. Pat. No.8,540,971, which is incorporated herein by reference. Suitable colorantsherein include pigments, dyes, lakes and agents imparting a particularluster or reflectivity such as pearling agents, for example. Specificexamples of colorants useful herein include talc; mica; magnesiumcarbonate; calcium carbonate; magnesium silicate; magnesium aluminumsilicate; silica; titanium dioxide; zinc oxide; red, yellow, brown andblack iron oxides; ferric ammonium ferrocyanide; manganese violet;ultramarine; titaniated mica; and bismuth oxychloride. One or morecolorants are optionally present in a total amount of about 0.001-20 wt% (e.g., about 0.01-10 wt % or about 0.1-5.0 wt %), for example, in thedisclosed oral care composition.

Additional components that can optionally be included in an oralcomposition herein include one or more enzymes (above), vitamins, andanti-adhesion agents, for example. Examples of vitamins useful hereininclude vitamin C, vitamin E, vitamin B5, and folic acid. Examples ofsuitable anti-adhesion agents include solbrol, ficin, and quorum-sensinginhibitors.

The present disclosure also concerns a method for increasing theviscosity of an aqueous composition. This method comprises contacting atleast one dextran compound as presently disclosed with the aqueouscomposition. The contacting step in this method results in increasingthe viscosity of the aqueous composition, in comparison to the viscosityof the aqueous composition before the contacting step.

An aqueous composition herein can be water (e.g., de-ionized water), anaqueous solution, or a hydrocolloid, for example. The viscosity of anaqueous composition before the contacting step, measured at about 20-25°C., can be about 0-10000 cPs (or any integer between 0-10000 cPs), forexample. Since the aqueous composition can be a hydrocolloid or the likein certain embodiments, it should be apparent that the method can beused to increase the viscosity of aqueous compositions that are alreadyviscous.

Contacting dextran herein with an aqueous composition increases theviscosity of the aqueous composition in certain embodiments. Thisincrease in viscosity can be an increase of at least about 1%, 10%,100%, 1000%, 100000%, or 1000000% (or any integer between 1% and1000000%), for example, compared to the viscosity of the aqueouscomposition before the contacting step. It should be apparent that verylarge percent increases in viscosity can be obtained with the disclosedmethod when the aqueous composition has little to no viscosity beforethe contacting step. An increase in viscosity can be determined, forexample, by comparing the viscosity of the aqueous composition obtainedby the method (i.e., after the contacting step) with the viscosity ofthe aqueous composition as it had existed before the method (i.e.,before the contacting step).

Contacting dextran herein with an aqueous composition increases theshear thinning behavior or shear thickening behavior of the aqueouscomposition in certain embodiments. Thus, dextran rheologically modifiesthe aqueous composition in these embodiments. The increase in shearthinning behavior or shear thickening behavior can be an increase of atleast about 1%, 10%, 100%, 1000%, 100000%, or 1000000% (or any integerbetween 1% and 1000000%), for example, compared to the shear thinningbehavior or shear thickening behavior of the aqueous composition beforethe contacting step. It should be apparent that very large percentincreases in rheologic modification can be obtained with the disclosedmethod when the aqueous composition has little to no rheologic behaviorbefore the contacting step.

The contacting step in a method for increasing the viscosity of anaqueous composition can be performed by mixing or dissolving any dextranas presently disclosed in the aqueous composition by any means known inthe art. For example, mixing or dissolving can be performed manually orwith a machine (e.g., industrial mixer or blender, orbital shaker, stirplate, homogenizer, sonicator, bead mill). Mixing or dissolving cancomprise a homogenization step in certain embodiments. Homogenization(as well as any other type of mixing) can be performed for about 5 to60, 5 to 30, 10 to 60, 10 to 30, 5 to 15, or 10 to 15 seconds (or anyinteger between 5 and 60 seconds), or longer periods of time asnecessary to mix dextran with the aqueous composition. A homogenizer canbe used at about 5000 to 30000 rpm, 10000 to 30000 rpm, 15000 to 30000rpm, 15000 to 25000 rpm, or 20000 rpm (or any integer between 5000 and30000 rpm), for example.

After a dextran herein is mixed with or dissolved into an aqueouscomposition, the resulting aqueous composition may be filtered, or maynot be filtered. For example, an aqueous composition prepared with ahomogenization step may or may not be filtered.

Certain embodiments of the above method can be used to prepare anaqueous composition disclosed herein, such as a food product (e.g., aconfectionery such as a candy filling), pharmaceutical product (e.g.,excipient), household product (e.g., laundry detergent, fabric softener,dishwasher detergent), personal care product (e.g., a water-containingdentifrice such as toothpaste), or industrial product.

The present disclosure also concerns a method of treating a material.This method comprises contacting a material with an aqueous compositioncomprising at least one dextran compound as disclosed herein.

A material contacted with an aqueous composition in a contacting methodherein can comprise a fabric in certain embodiments. A fabric herein cancomprise natural fibers, synthetic fibers, semi-synthetic fibers, or anycombination thereof. A semi-synthetic fiber herein is produced usingnaturally occurring material that has been chemically derivatized, anexample of which is rayon. Non-limiting examples of fabric types hereininclude fabrics made of (i) cellulosic fibers such as cotton (e.g.,broadcloth, canvas, chambray, chenille, chintz, corduroy, cretonne,damask, denim, flannel, gingham, jacquard, knit, matelassé, oxford,percale, poplin, plissé, sateen, seersucker, sheers, terry cloth, twill,velvet), rayon (e.g., viscose, modal, lyocell), linen, and Tencel®; (ii)proteinaceous fibers such as silk, wool and related mammalian fibers;(iii) synthetic fibers such as polyester, acrylic, nylon, and the like;(iv) long vegetable fibers from jute, flax, ramie, coir, kapok, sisal,henequen, abaca, hemp and sunn; and (v) any combination of a fabric of(i)-(iv). Fabric comprising a combination of fiber types (e.g., naturaland synthetic) include those with both a cotton fiber and polyester, forexample. Materials/articles containing one or more fabrics hereininclude, for example, clothing, curtains, drapes, upholstery, carpeting,bed linens, bath linens, tablecloths, sleeping bags, tents, carinteriors, etc. Other materials comprising natural and/or syntheticfibers include, for example, non-woven fabrics, paddings, paper, andfoams.

An aqueous composition that is contacted with a fabric can be, forexample, a fabric care composition (e.g., laundry detergent, fabricsoftener). Thus, a treatment method in certain embodiments can beconsidered a fabric care method or laundry method if employing a fabriccare composition therein. A fabric care composition herein iscontemplated to effect one or more of the following fabric care benefits(i.e., surface substantive effects): wrinkle removal, wrinkle reduction,wrinkle resistance, fabric wear reduction, fabric wear resistance,fabric pilling reduction, extended fabric life, fabric colormaintenance, fabric color fading reduction, reduced dye transfer, fabriccolor restoration, fabric soiling reduction, fabric soil release, fabricshape retention, fabric smoothness enhancement, anti-redeposition ofsoil on fabric, anti-greying of laundry, improved fabric hand/handle,and/or fabric shrinkage reduction.

Examples of conditions (e.g., time, temperature, wash/rinse volumes) forconducting a fabric care method or laundry method herein are disclosedin WO1997/003161 and U.S. Pat. Nos. 4,794,661, 4,580,421 and 5,945,394,which are incorporated herein by reference. In other examples, amaterial comprising fabric can be contacted with an aqueous compositionherein: (i) for at least about 5, 10, 20, 30, 40, 50, 60, 70, 80, 90,100, 110, or 120 minutes; (ii) at a temperature of at least about 10,15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, or 95°C. (e.g., for laundry wash or rinse: a “cold” temperature of about15-30° C., a “warm” temperature of about 30-50° C., a “hot” temperatureof about 50-95° C.); (iii) at a pH of about 2, 3, 4, 5, 6, 7, 8, 9, 10,11, or 12 (e.g., pH range of about 2-12, or about 3-11); (iv) at a salt(e.g., NaCl) concentration of at least about 0.5, 1.0, 1.5, 2.0, 2.5,3.0, 3.5, or 4.0 wt %; or any combination of (i)-(iv).

The contacting step in a fabric care method or laundry method cancomprise any of washing, soaking, and/or rinsing steps, for example.Contacting a material or fabric in still further embodiments can beperformed by any means known in the art, such as dissolving, mixing,shaking, spraying, treating, immersing, flushing, pouring on or in,combining, painting, coating, applying, affixing to, and/orcommunicating an effective amount of a dextran compound herein with thefabric or material. In still further embodiments, contacting may be usedto treat a fabric to provide a surface substantive effect. As usedherein, the term “fabric hand” or “handle” refers to a person's tactilesensory response towards fabric which may be physical, physiological,psychological, social or any combination thereof. In one embodiment, thefabric hand may be measured using a PhabrOmeter® System for measuringrelative hand value (available from Nu Cybertek, Inc. Davis, Calif.)(American Association of Textile Chemists and Colorists (AATCC testmethod “202-2012, Relative Hand Value of Textiles: InstrumentalMethod”)).

In certain embodiments of treating a material comprising fabric, adextran compound component(s) of the aqueous composition adsorbs to thefabric. This feature is believed to render dextran compounds hereinuseful as anti-redeposition agents and/or anti-greying agents in fabriccare compositions disclosed (in addition to their viscosity-modifyingeffect). An anti-redeposition agent or anti-greying agent herein helpskeep soil from redepositing onto clothing in wash water after the soilhas been removed. It is further contemplated that adsorption of one ormore dextran compounds herein to a fabric enhances mechanical propertiesof the fabric.

Adsorption of a dextran compound to a fabric herein can be measuredusing a colorimetric technique (e.g., Dubois et al., 1956, Anal. Chem.28:350-356; Zemlji

et al., 2006, Lenzinger Berichte 85:68-76; both incorporated herein byreference), for example, or any other method known in the art.

Other materials that can be contacted in the above treatment methodinclude surfaces that can be treated with a dish detergent (e.g.,automatic dishwashing detergent or hand dish detergent). Examples ofsuch materials include surfaces of dishes, glasses, pots, pans, bakingdishes, utensils and flatware made from ceramic material, china, metal,glass, plastic (e.g., polyethylene, polypropylene, polystyrene, etc.)and wood (collectively referred to herein as “tableware”). Thus, thetreatment method in certain embodiments can be considered a dishwashingmethod or tableware washing method, for example. Examples of conditions(e.g., time, temperature, wash volume) for conducting a dishwashing ortableware washing method herein are disclosed in U.S. Pat. No.8,575,083, which is incorporated herein by reference. In other examples,a tableware article can be contacted with an aqueous composition hereinunder a suitable set of conditions such as any of those disclosed abovewith regard to contacting a fabric-comprising material.

Other materials that can be contacted in the above treatment methodinclude oral surfaces such as any soft or hard surface within the oralcavity including surfaces of the tongue, hard and soft palate, buccalmucosa, gums and dental surfaces (e.g., natural tooth or a hard surfaceof artificial dentition such as a crown, cap, filling, bridge, denture,or dental implant). Thus, a treatment method in certain embodiments canbe considered an oral care method or dental care method, for example.Conditions (e.g., time, temperature) for contacting an oral surface withan aqueous composition herein should be suitable for the intendedpurpose of making such contact. Other surfaces that can be contacted ina treatment method also include a surface of the integumentary systemsuch as skin, hair or nails.

Thus, certain embodiments of the present disclosure concern material(e.g., fabric) that comprises a dextran compound herein. Such materialcan be produced following a material treatment method as disclosedherein, for example. A material may comprise a dextran compound incertain embodiments if the compound is adsorbed to, or otherwise incontact with, the surface of the material.

Certain embodiments of a method of treating a material herein furthercomprise a drying step, in which a material is dried after beingcontacted with the aqueous composition. A drying step can be performeddirectly after the contacting step, or following one or more additionalsteps that might follow the contacting step (e.g., drying of a fabricafter being rinsed, in water for example, following a wash in an aqueouscomposition herein). Drying can be performed by any of several meansknown in the art, such as air drying (e.g., ˜20-25° C.), or at atemperature of at least about 30, 40, 50, 60, 70, 80, 90, 100, 120, 140,160, 170, 175, 180, or 200° C., for example. A material that has beendried herein typically has less than 3, 2, 1, 0.5, or 0.1 wt % watercomprised therein. Fabric is a preferred material for conducting anoptional drying step.

An aqueous composition used in a treatment method herein can be anyaqueous composition disclosed herein, such as in the above embodimentsor in the below Examples. Thus, the dextran component(s) of an aqueouscomposition can be any as disclosed herein. Examples of aqueouscompositions include detergents (e.g., laundry detergent or dishdetergent) and water-containing dentifrices such as toothpaste.

The present disclosure also concerns an enzymatic reaction comprisingwater, sucrose and a glucosyltransferase enzyme comprising, orconsisting of, an amino acid sequence that is at least 90% identical toSEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:9, SEQ ID NO:13, or SEQID NO:17. The glucosyltransferase enzyme synthesizes dextran aspresently disclosed. Significantly, dextran synthesized in this gtfreaction exhibits high viscosity in aqueous compositions, even atrelatively low concentrations of the dextran. It is believed that thishigh viscosity profile is unique in comparison to viscosity profiles ofpreviously disclosed dextran polymers.

Dextran synthesized in an enzymatic reaction herein can be ascharacterized (e.g., molecular weight, linkage and branching profile) inthe above disclosure regarding dextran as produced by aglucosyltransferase enzyme. A glucosyltransferase enzyme in an enzymaticreaction herein can be as characterized in the above disclosureregarding dextran as produced by a glucosyltransferase enzyme.

One or more different glucosyltransferase enzymes may be used in anenzymatic reaction herein. A single glucosyltransferase enzyme (e.g.,gtf 0768) is used in some cases, as opposed to situations in whichmultiple enzymes may be present (e.g., a bacterial or yeastfermentation). An enzymatic reaction can be as characterized (e.g.,initial sucrose concentration and sucrose type, pH, temperature, time)in the above disclosure regarding dextran as produced by aglucosyltransferase enzyme. Also, any features presently disclosed of amethod of producing dextran can apply to a glucosyltransferase reaction.

The present disclosure also concerns a method for producing dextrancomprising the step of contacting at least water, sucrose, and aglucosyltransferase enzyme comprising an amino acid sequence that is atleast 90% identical to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:5, SEQ IDNO:9, SEQ ID NO:13, or SEQ ID NO:17. This contacting step results inproduction of dextran as presently disclosed. Dextran produced in thecontacting step can optionally be isolated.

Dextran synthesized in a synthesis method herein can be as characterized(e.g., molecular weight, linkage and branching profile) in the abovedisclosure regarding dextran as produced by a glucosyltransferaseenzyme. A glucosyltransferase enzyme in a synthesis method herein can beas characterized in the above disclosure regarding dextran as producedby a glucosyltransferase enzyme. Any features of an enzymatic reactionas disclosed above can apply to the instant synthesis method.

The contacting step in a method herein of producing dextran comprisesproviding an enzymatic reaction comprising water, sucrose and anyglucosyltransferase enzyme disclosed herein. The contacting step of thedisclosed method can be performed in any number of ways. For example,the desired amount of sucrose can first be dissolved in water(optionally, other components may also be added at this stage ofpreparation, such as buffer components), followed by addition of one ormore glucosyltransferase enzymes. The solution may be kept still, oragitated via stirring or orbital shaking, for example.

The reaction can be, and typically is, cell-free. Thus, a dextran hereinis not isolated from a cell, such as a bacteria (e.g., L.mesenteroides), in some aspects.

Completion of a glucosyltransferase reaction in certain embodiments canbe gauged, for example, by determining whether reaction viscosity is nolonger increasing and/or by measuring the amount of sucrose left in thereaction (residual sucrose), where a percent sucrose consumption of overabout 90% can indicate reaction completion. Typically, a reaction of thedisclosed process can take about 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 18, 24,30, 36, 48, 60, 72, 84, or 96 hours to complete. Reaction time maydepend, for example, on certain parameters such as the amount of sucroseand glucosyltransferase enzyme used in the reaction.

The yield of dextran produced in a glucosyltransferase reaction incertain embodiments can be about, or at least about, 10%, 15%, 20%, 25%,30%, 35%, 40%, or 45%, based on the weight of the sucrose used in thereaction.

Dextran produced in the disclosed method may optionally be isolated. Forexample, dextran may be precipitated with alcohol (e.g., 90-100%methanol, ethanol, or isopropanol) and then separated from thesupernatant, which may comprise water, fructose, and optionally one ormore of residual sucrose and byproduct (e.g., glucose; leucrose andother soluble oligosaccharides). Such separation can be bycentrifugation or filtration, for example. Precipitated dextran canoptionally be washed one or more times (e.g., 2-4 times; 2, 3, 4 or moretimes) with alcohol (e.g., 70-100%, or at least 70%, 80%, 90%, 95%, or100% methanol, ethanol, or isopropanol). In other examples, dextranisolation can comprise using an ultrafiltration and/or dialysistechnique (i.e., a molecular weight cut-off technique), such asdisclosed in U.S. Patent Appl. Publ. No. 2014/0142294 and U.S. Pat. No.6,977,249, which are incorporated herein by reference. Measurements ofcertain dextran features herein (e.g., linkage profile, molecularweight) can be made with dextran isolated as above, if desired.

A dextran synthesis method herein is believed to be useful for producingdextran with increased or decreased viscosity, depending on the amountof sucrose used in the method. In general, the lower the sucroseconcentration used in a glucosyltransferase reaction, the higher theviscosity of the dextran product, and vice versa. Any sucroseconcentration disclosed herein can be used in a glucosyltransferasereaction, where the dextran product of the reaction has a viscosity thatis greater than that of a dextran product produced in a reactioncomprising a higher sucrose concentration, and vice versa. In certainaspects, any viscosity disclosed herein can be used to characterizeembodiments of this method, and an increase in viscosity can be at leastabout 2-, 3-, 4-, 5-, 6-, 7-, 8-, 9-, 10-, 20-, 50-, 100-, 150-, 200-,or 250-fold higher. A glucosyltransferase enzyme in certain embodimentsof this method can be gtf 0768 (comprising SEQ ID NO:1 or relatedsequences).

Non-limiting examples of compositions and methods disclosed hereininclude:

-   1. A composition comprising dextran, wherein the dextran comprises:-   (i) about 87-93 wt % glucose linked at positions 1 and 6;-   (ii) about 0.1-1.2 wt % glucose linked at positions 1 and 3;-   (iii) about 0.1-0.7 wt % glucose linked at positions 1 and 4;-   (iv) about 7.7-8.6 wt % glucose linked at positions 1, 3 and 6; and-   (v) about 0.4-1.7 wt % glucose linked at:    -   (a) positions 1, 2 and 6, or    -   (b) positions 1, 4 and 6;        wherein the weight-average molecular weight (Mw) of the dextran        is about 50-200 million Daltons, the z-average radius of        gyration of the dextran is about 200-280 nm, and the dextran        optionally is not a product of a Leuconostoc mesenteroides        glucosyltransferase enzyme.-   2. The composition of embodiment 1, wherein the dextran comprises:-   (i) about 89.5-90.5 wt % glucose linked at positions 1 and 6;-   (ii) about 0.4-0.9 wt % glucose linked at positions 1 and 3;-   (iii) about 0.3-0.5 wt % glucose linked at positions 1 and 4;-   (iv) about 8.0-8.3 wt % glucose linked at positions 1, 3 and 6; and-   (v) about 0.7-1.4 wt % glucose linked at:    -   (a) positions 1, 2 and 6, or    -   (b) positions 1, 4 and 6.-   3. The composition of embodiment 1 or 2, wherein the dextran    comprises chains linked together within a branching structure,    wherein the chains are similar in length and comprise substantially    alpha-1,6-glucosidic linkages.-   4. The composition of embodiment 1, 2, or 3, wherein the average    length of the chains is about 10-50 monomeric units.-   5. The composition of embodiment 1, 2, 3, or 4, wherein the dextran    is a product of a glucosyltransferase enzyme comprising an amino    acid sequence that is at least 90% identical to SEQ ID NO:1, SEQ ID    NO:2, SEQ ID NO:5, SEQ ID NO:9, SEQ ID NO:13, or SEQ ID NO:17.-   6. The composition of embodiment 1, 2, 3, 4, or 5, wherein the    composition is an aqueous composition having a viscosity of at least    about 25 cPs.-   7. The composition of embodiment 1, 2, 3, 4, 5, or 6, wherein the Mw    of the dextran is about 80-120 million Daltons.-   8. The composition of embodiment 1, 2, 3, 4, 5, 6, or 7, wherein the    z-average radius of gyration of the dextran is about 230-250 nm.-   9. The composition of embodiment 1, 2, 3, 4, 5, 6, 7, or 8, wherein    the composition is in the form of a food product, personal care    product, pharmaceutical product, household product, or industrial    product.-   10. The composition of embodiment 9, wherein the composition is in    the form of a confectionery.-   11. A method for increasing the viscosity of an aqueous composition,    the method comprising: contacting dextran according to any of    embodiments 1-8 with the aqueous composition, wherein the viscosity    of the aqueous composition is increased by the dextran compared to    the viscosity of the aqueous composition before the contacting step.-   12. A method of treating a material, the method comprising:    contacting a material with an aqueous composition comprising dextran    according to any of embodiments 1-8.-   13. An enzymatic reaction comprising water, sucrose and a    glucosyltransferase enzyme comprising an amino acid sequence that is    at least 90% identical to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:5, SEQ    ID NO:9, SEQ ID NO:13, or SEQ ID NO:17, wherein the    glucosyltransferase enzyme synthesizes dextran according to any of    embodiments 1-8.-   14. A method for producing dextran, the method comprising:-   a) contacting at least water, sucrose, and a glucosyltransferase    enzyme comprising an amino acid sequence that is at least 90%    identical to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:9, SEQ    ID NO:13, or SEQ ID NO:17, whereby dextran according to any of    embodiments 1-8 is produced; and-   b) optionally, isolating the dextran produced in step (a).-   15. The method of embodiment 14, wherein the viscosity of the    dextran produced in the method is increased by decreasing the amount    of sucrose in step (a).

EXAMPLES

The present disclosure is further defined in Examples 1-6 and 8-11. Itshould be understood that these Examples, while indicating certainpreferred aspects of the disclosure, are given by way of illustrationonly. From the above discussion and these Examples, one skilled in theart can ascertain the essential characteristics of this disclosure, andwithout departing from the spirit and scope thereof, can make variouschanges and modifications to adapt the disclosure to various uses andconditions.

General Methods

Cloning and Expression of Glucosyltransferase Enzymes in Bacillussubtilis

Each glucosyltransferase used in Examples 3-6 was prepared as follows.

A plasmid encoding the gtf enzyme (pZZHB582, pZZHB583, pZZHB584, orpZZHB585, which allow for gtf expression and secretion from B. subtilis;see FIGS. 2A-D) was amplified using Illustra TempliPhi® 100Amplification Kit (GE Healthcare Life Sciences, NJ). Competent B.subtilis cells (ΔspoIIE, ΔaprE, ΔnprE, degUHy32, ΔscoC, ΔnprB, Δvpr,Δepr, ΔwprA, Δmpr, ΔispA, Δbpr) were transformed with the amplificationproduct. Cells were plated on Luria Agar plates supplemented with 5 ppmchloramphenicol. Colonies from the transformation plate were inoculatedinto 5 mL LB medium and incubated at 37° C. overnight. Aliquots (25-50μL) from each culture were then transferred to 250-mL shake flaskscontaining 30 mL of Grant's II Medium supplemented with 5 ppmchloramphenicol and incubated at 30° C. with shaking (280 rpm) for 24hours. Cells were harvested by centrifugation at 14000 rpm for 1 hour.Supernatants were analyzed by SDS-PAGE for secreted gtf product andfurther dialyzed three times against a solution containing 20 mM Tris,pH 7.5 for a total of 20 hours. Dialyzed samples were aliquoted at 25 mLper 50-mL conical centrifuge tube, and the tubes were placed at an angleat −80° C. for about 1 hour. Once the samples were frozen, the tube lidwas removed and replaced with PARAFILM that was pierced 5-10 times witha high-gauge needle. The PARAFILM-covered frozen samples werelyophilized in a FreeZone® Freeze Dry System (Labconco Corp., KansasCity, Mo.) according to the manufacturer's instruction.

Stock Solutions of Glucosyltransferase Enzymes

An enzyme stock solution was made for each gtf by adding 10 mL ofmolecular grade H₂O into each 50-mL conical centrifuge tube containinglyophilized enzyme powder.

Example 1 Expression of a Glucosyltransferase (0768) in E. coli andProduction of Active Crude Enzyme Lysate

This Example describes expression of a mature glucosyltransferase (gtf)enzyme in E. coli. Crude cell lysate of an E. coli expression strain wasproduced and showed gel product-forming activity in the presence ofsucrose.

A putative YG repeat-containing hydrolase (categorized in GENBANK underGI number 339480768, but now having GI number 497964659) with 1484 aminoacids was identified from Leuconostoc pseudomesenteroides strainKCTC3652 by whole genome shotgun sequencing. This putativeglucosyltransferase (designated herein as gtf 0768) belongs to the GH70family of glycosyl hydrolases containing a glucan-binding domain. TheN-terminal 37 amino acid segment of gtf 0768 was deduced as the signalpeptide of the enzyme by the SIGNALP 4.0 program (Petersen et al.,Nature Methods 8:785-786). The mature form of gtf 0768 is represented bySEQ ID NO:1.

To construct a plasmid for bacterial expression of gtf 0768, a DNAsequence encoding a mature form of the gtf without the signal peptidewas synthesized by GenScript USA Inc. (Piscataway, N.J.). Thesynthesized sequence was subcloned into the NheI and HindIII sites ofthe pET23D+ vector (NOVAGEN®; Merck KGaA, Darmstadt, Germany). The 0768gtf (SEQ ID NO:2) encoded by this construct included a start methionineand 3 additional amino acids (Ala-Ser-Ala) at the N-terminus, and 6histidine residues at the C-terminus, compared to the wild type mature(predicted) form of gtf 0768 (SEQ ID NO:1) (i.e., SEQ ID NO:1 iscomprised in SEQ ID NO:2). The plasmid construct was sequence-confirmedand transformed into E. coli BL21 DE3 host cells with ampicillinselection, resulting in expression strain EC0052.

Cells of EC0052 and a control strain containing only empty pET23D+vector were grown in LB medium with 100 μg/mL ampicillin to OD₆₀₀ ˜0.5,and then induced with 1 mM IPTG at 37° C. for 3 hours or alternativelyinduced at 23° C. overnight. Following this induction period, cells werecollected by centrifugation at 4000×g for 10 min and resuspended in PBSbuffer pH 6.8. The cells were then lysed by passing through a FrenchPress at 14,000 psi (˜96.53 MPa) twice, after which cell debris waspelleted by centrifugation at 15,000×g for 20 min. The supernatants ofeach crude cell lysate were aliquoted and frozen at −80° C.

The activity of crude cell lysate from EC0052 cells was checked byreaction with sucrose. A control reaction was set up similarly usingcell lysate prepared from cells containing the empty vector. Eachsucrose reaction was set up using 10% (v/v) of cell lysate with 100 g/Lsucrose, 10 mM sodium citrate pH 5, and 1 mM CaCl₂. After incubation ofthe reactions at 37° C. for a few hours, a gel-like product, believed tobe a dextran, was formed in the tube in which EC0052 cell lysate hadbeen added. No gel-like product was formed in the control reaction. HPLCanalysis confirmed that sucrose was consumed in the reaction containingEC0052 cell lysate, and not in the control reaction. This resultsuggested that the EC0052 crude cell lysate expressed active gtf 0768enzyme, and that this gtf produced a dextran product having highviscosity.

Thus, reactions comprising water, sucrose and an enzyme comprising SEQID NO:1 synthesized a gelling product, believed to be a dextran. Thisresult demonstrated that gtf 0768 likely has glucosyltransferaseactivity.

Example 2 Reaction of Sucrose with Gtf 0768 and Analysis of a GellingDextran Reaction Product

This Example describes additional reactions comprising water, sucroseand gtf 0768, supplementing the results provided in Example 1. Also,this Example provides glycosidic linkage analysis of the gelling productsynthesized by gtf 0768, showing that this product is a type of dextran.

Reagents for preparing gtf reactions:

-   -   Sucrose (Sigma Prod. No. S-9378).    -   Sodium phosphate buffer stock (200 mM) (pH 5.5): prepare 250 mL        in water using sodium phosphate monobasic monohydrate (Sigma        Prod. No. S9638) and sodium phosphate dibasic heptahydrate        (Sigma Prod. No. S9390), accordingly.    -   Gtf 0768 enzyme solution (cell lysate as prepared in Example 1).

Conditions of three gtf reactions:

A 1000-mL reaction was prepared containing 2.72 g of sodium phosphatebuffer stock (pH 5.5), 100 g/L sucrose, and 2 mL of gtf 0768 enzymesolution. The reaction was stirred at 26° C. for 20 hours, and becameviscous. The gtf enzyme was deactivated by heating the reaction at 80°C. for 10 minutes. The deactivated viscous reaction was then mixed with3 liters of 100% methanol to precipitate the viscous product. A whiteprecipitate was formed, which was then filtered, followed by four washeswith 120 ml of 100% methanol. The solid product was dried at roomtemperature under vacuum in an oven for 72 hours.

A 725-mL reaction was prepared containing 1.97 g of sodium phosphatebuffer, 300 g/L sucrose, and 1.45 mL of gtf 0768 enzyme solution. Thereaction was stirred at 26° C. for 20 hours, and became viscous. The gtfenzyme was deactivated by adding methanol to the reaction mixture. Thedeactivated reaction was then mixed with 3 liters of 100% methanol toprecipitate the viscous product. A white precipitate was formed, whichwas then filtered, followed by four washes with 120 mL of 100% methanol.The solid product was dried at room temperature under vacuum in an ovenfor 72 hours.

A 200-mL reaction was prepared containing 0.544 g of sodium phosphatebuffer, 400 g/L sucrose, and 0.4 mL of gtf 0768 enzyme solution. Thereaction was stirred at 26° C. for 20 hours, and became viscous. The gtfenzyme was deactivated by adding methanol to the reaction mixture. Thedeactivated reaction was then mixed with 3 liters of 100% methanol toprecipitate the viscous product. A white precipitate was formed, whichwas then filtered, followed by four washes with 120 mL of 100% methanol.The solid product was dried at room temperature under vacuum in an ovenfor 72 hours.

A 200-mL reaction was prepared containing 0.544 g of sodium phosphatebuffer, 800 g/L sucrose, and 0.4 mL of gtf 0768 enzyme solution. Thereaction was stirred at 26° C. for 20 hours, and became viscous. The gtfenzyme was deactivated by adding methanol to the reaction mixture. Thedeactivated reaction was then mixed with 3 liters of 100% methanol toprecipitate the viscous product. A white precipitate was formed, whichwas then filtered, followed by four washes with 120 ml of 100% methanol.The solid product was dried at room temperature under vacuum in an ovenfor 72 hours.

Samples (100 μL) of each reaction were taken at 0, 2, 4, and 18 hours,respectively. The gtf enzyme was deactivated in each sample by heatingat 80° C. for 10 minutes. Each sample was then diluted 10-fold withwater and centrifuged at 14,000 rpm for 5 minutes, after which 200 μl ofsupernatant was used for HPLC analysis to measure sucrose consumptionduring the reaction. The following HPLC conditions were applied foranalyzing each sample: column (AMINEX HPX-87C carbohydrate column,300×7.8 mm, Bio-Rad, No. 125-0095), eluent (water), flow rate (0.6mL/min), temperature (85° C.), refractive index detector. HPLC analysisof the samples indicated substantial sucrose consumption during the 0768gtf reaction (FIG. 1, reaction comprising 100 g/L sucrose) (this sucroseconsumption occurred significantly faster than the sucrose consumptionobserved in a reaction using a dextran sucrase obtained from acommercial source—refer to Example 7).

HPLC was also used to analyze other products of the reaction comprising100 g/L sucrose. Polymer yield was back-calculated by subtracting theamount of all other saccharides left in the reaction from the amount ofthe starting sucrose. The back-calculated number was consistent with theviscous product dry weight analysis. Sucrose, leucrose, glucose andfructose were quantified by HPLC with an HPX-87C column (HPLC conditionsas described above). DP2-7 disaccharides were quantified by HPLC withthe following conditions: column (AMINEX HPX-42A carbohydrate column,300×7.8 mm, Bio-Rad, No. 125-0097), eluent (water), flow rate (0.6mL/min), temperature (85° C.), refractive index detector. These HPLCanalyses indicated that the glucosyl-containing saccharide products ofthe 0768 gtf reaction consisted of 91% polymer product, 1% glucose, 6.5%leucrose, and 1.5% DP2-7 oligosaccharides.

The glycosidic linkage profile of the gelling polymer product of thereaction comprising 100 g/L sucrose was determined by ¹³C NMR. Drypolymer (25-30 mg) as prepared above was dissolved in 1 mL of deuteratedDMSO containing 3 wt % LiCl with stirring at 50° C. Using a glass pipet,0.8 mL of the preparation was transferred into a 5-mm NMR tube. Aquantitative ¹³C NMR spectrum was acquired using a Bruker Avance(Billerica, Mass.) 500 MHz NMR spectrometer equipped with a CPDuIcryoprobe, at a spectral frequency of 125.76 MHz, using a spectralwindow of 26041.7 Hz. An inverse-gated decoupling pulse sequence usingwaltz decoupling was used with an acquisition time of 0.629 second, aninter-pulse delay of 5 seconds, and 6000 pulses. The time domain datawere transformed using an exponential multiplication of 2.0 Hz.

The NMR results indicated that the gelling polymer product comprisedabout 90% alpha-1,6-glucosidic linkages, about 4-5% alpha-1,3-glucosidiclinkages, and about 5-6% alpha-1,4 and -1,2 glucosidic linkages. Themain chain(s) of the polymer product appeared to mostly comprisealpha-1,6-glucosidic linkages, but also a very small amount of alpha-1,3and -1,4 glucosidic linkages. Other alpha-1,3 and -1,4 glucosidiclinkages, and all of the alpha-1,2-glucosidic linkages, appeared to bein branches off the main chain(s). The gelling product thus appears tobe a gelling dextran.

A different protocol (not the above ¹³C NMR procedure) is presentlyrecommended herein for determining the linkage profile of dextranproduced by gtf 0768. This protocol is disclosed below in Example 9,indicating a linkage profile similar to that disclosed in this Example.

The number-average molecular weight (Mn) and weight-average molecularweight (Mw) of the gelling dextran product of the reaction comprising100 g/L sucrose was determined by size-exclusion chromatography (SEC).Dry polymer as prepared above was dissolved in DMAc and 5% LiCl (0.5mg/mL) with shaking overnight at 100° C. The chromatographic system usedwas an Alliance™ 2695 separation module from Waters Corporation(Milford, Mass.) coupled with three on-line detectors: a differentialrefractometer 2410 from Waters, a Heleos™ 8+ multiangle light scatteringphotometer from Wyatt Technologies (Santa Barbara, Calif.), and aViscoStar™ differential capillary viscometer from Wyatt. Columns usedfor SEC were four styrene-divinyl benzene columns from Shodex (Japan)and two linear KD-806M, KD-802 and KD-801 columns to improve resolutionat the low molecular weight region of a polymer distribution. The mobilephase was DMAc with 0.11% LiCl. The chromatographic conditions used were50° C. in the column and detector compartments, 40° C. in the sample andinjector compartment, a flow rate of 0.5 mL/min, and an injection volumeof 100 μL. The software packages used for data reduction were Empower™version 3 from Waters (calibration with broad glucan polymer standard)and Astra® version 6 from Wyatt (triple detection method with columncalibration). It was determined from this procedure that the gellingdextran product had an M_(n) of 2229400 and an Mw of 5365700.

A different protocol (not the above SEC procedure) is presentlyrecommended herein for determining the molecular weight of dextranproduced by gtf 0768. This protocol is disclosed below in Example 9,indicating a molecular weight more than one order of magnitude greaterthan the molecular weight disclosed in this Example.

Thus, reactions comprising water, sucrose and an enzyme comprising SEQID NO:1 synthesized a gelling dextran product, as determined by theproduct's predominant alpha-1,6 glucosidic linkage profile. Example 8below discloses comparing the viscosity of this product versus theviscosities of certain commercially available dextrans. Example 9discloses further production of dextran with a gtf enzyme comprising SEQID NO:1, along with yield, molecular weight, and linkage analysis of thedextran.

Example 3 Expression of a Glucosyltransferase (2919) and Use Thereof toProduce a Gelling Dextran Product

This Example describes expression of a mature Weissella cibariaglucosyltransferase (gtf) enzyme in B. subtilis. Also, this Exampleshows that this enzyme produces a gelling product, likely a dextran,when used is a reaction containing water and sucrose.

A glucosyltransferase gene, WciGtf1, was identified from Weissellacibaria KACC 11862. The nucleic acid sequence of this gene (positions23315 to 27661 of GENBANK Accession No. NZ_AEKT01000035.1) is set forthin SEQ ID NO:3 and encodes the protein sequence of SEQ ID NO:4 (GENBANKAccession No. ZP_08417432). At the N-terminus of the WciGtf1 protein(SEQ ID NO:4) is a signal peptide of 26 amino acids, as predicted by theSIGNALP 4.0 program (Petersen et al., Nature Methods 8:785-786). Thisindicates that WciGtf1 (SEQ ID NO:4) is a secreted protein. The mature,secreted form of the WciGtf1 protein is herein referred to as 2919 gtf,and is set forth in SEQ ID NO:5.

The nucleotide sequence encoding 2919 gtf was optimized for expressionin B. subtilis. The optimized sequence (SEQ ID NO:6) was synthesized byGeneray (Shanghai, China), and inserted into plasmid p2JM103BBI(Vogtentanz et al., Protein Expr. Purif. 55:40-52), resulting in plasmidpZZHB583 (FIG. 2A). Plasmid pZZHB583 contains an aprE promoter operablylinked to a sequence encoding (i) an aprE signal sequence used to directheterologous protein (2919 gtf in this case) secretion in B. subtilis,(ii) Ala-Gly-Lys to facilitate the secretion, and (iii) 2919 gtf (SEQ IDNO:5) (i-iii are fused together in the amino-to-carboxy direction).

Plasmid pZZHB583 was transformed into B. subtilis cells for 2919 gtfexpression and purification (see General Methods).

The activity of 2919 gtf (SEQ ID NO:5) was determined in a 250-mLreaction at room temperature comprising 100 g/L sucrose, 20 mM sodiumphosphate buffer (pH 5.5), and 6.25 mL of enzyme stock. The reaction wascarried out at room temperature with shaking (150 rpm) for 48 hours.

Samples (100 μL) were taken from the reaction at 0, 1, 3, 5, 24, and 48hour time points, respectively. Enzyme was deactivated by heating eachsample at 80° C. for 10 minutes. Samples were diluted 10-fold with waterand centrifuged at 14000 rpm for 5 minutes. Supernatant (200 μL) wasused for HPLC analysis.

The concentrations of leucrose, glucose, and fructose in the gtfreaction were determined using HPLC, which was performed with an Agilent1260 chromatography system equipped with an AMINEX HPX-87C column(300×7.8 mm) placed in a thermostatted column compartment at 85° C., anda refractive index detector. HPLC elution was carried out with Milli-Q®water at 0.6 mL/min. Sucrose, leucrose, glucose, and fructose wereidentified by comparison with corresponding standards. Theirconcentrations were calculated based on a peak area standard curves.Sucrose was consumed almost completely by the end of the reaction. Asidefrom a viscous dextran product, 2919 gtf (SEQ ID NO:5) produced mostlyfructose (˜50%), and small amounts of leucrose (˜5%) and glucose (˜1%).

The concentration of oligosaccharides (DP2-DP7) in the gtf reaction wasdetermined by HPLC analysis, which was performed with an Agilent 1260chromatography system equipped with an AMINEX HPX-42A column (300×7.8mm) placed in a thermostatted column compartment at 85° C., and arefractive index detector. HPLC elution was carried out with Milli-Q®water at 0.6 mL/min. Formation of oligosaccharides was identified bycomparison with corresponding standards. The concentration of theoligosaccharides was calculated based on standard curves from peak area.2919 gtf (SEQ ID NO:5) produced a small amount of DP2-DP7oligosaccharides (˜3%) by the end of the reaction.

Thus, reactions comprising water, sucrose and an enzyme comprising SEQID NO:5 synthesized a gelling product, which is believed to be a dextranpolymer. Experimental results demonstrated that gtf 2919 likely hasglucosyltransferase activity.

Example 4 Expression of a Glucosyltransferase (2918) and Use Thereof toProduce a Gelling Dextran Product

This Example describes expression of a mature Lactobacillus fermentumglucosyltransferase (gtf) enzyme in B. subtilis. Also, this Exampleshows that this enzyme produces a gelling product, likely a dextran,when used is a reaction containing water and sucrose.

A glucosyltransferase gene, LfeGtf1, was identified from Lactobacillusfermentum. The nucleic acid sequence of this gene (positions 618 to 5009of GENBANK Accession No. AY697433.1) is set forth in SEQ ID NO:7 andencodes the protein sequence of SEQ ID NO:8 (GENBANK Accession No.AAU08008). At the N-terminus of the LfeGtf1 protein (SEQ ID NO:8) is asignal peptide of 37 amino acids, as predicted by the SIGNALP 4.0program. This indicates that LfeGtf1 (SEQ ID NO:8) is a secretedprotein. The mature, secreted form of the LfeGtf1 protein is hereinreferred to as 2918 gtf, and is set forth in SEQ ID NO:9.

The nucleotide sequence encoding 2918 gtf was optimized for expressionin B. subtilis. The optimized sequence (SEQ ID NO:10) was synthesized byGeneray (Shanghai, China), and inserted into plasmid p2JM103BBI,resulting in plasmid pZZHB582 (FIG. 2B). Plasmid pZZHB582 contains anaprE promoter operably linked to a sequence encoding (i) an aprE signalsequence used to direct heterologous protein (2918 gtf in this case)secretion in B. subtilis, (ii) Ala-Gly-Lys to facilitate the secretion,and (iii) 2918 gtf (SEQ ID NO:9) (i-iii are fused together in theamino-to-carboxy direction).

Plasmid pZZHB582 was transformed into B. subtilis cells for 2918 gtfexpression and purification (see General Methods).

The activity of 2918 gtf (SEQ ID NO:9) was determined in a 250-mLreaction at room temperature comprising 100 g/L sucrose, 20 mM sodiumphosphate buffer (pH 5.5), and 6.25 mL of enzyme stock. The reaction wascarried out at room temperature with shaking (150 rpm) for 6 days.

Samples (100 μL) were taken from the reaction at 0, 1, 3, 5, 24, 48 and144 hour time points, respectively. Enzyme was deactivated by heatingeach sample at 80° C. for 10 minutes. Samples were diluted 10-fold withwater and centrifuged at 14000 rpm for 5 minutes. Supernatant (200 μL)was used for HPLC analysis.

The concentrations of sucrose, leucrose, glucose, fructose andoligosaccharides (DP2-DP7) in the gtf reaction were determined usingHPLC procedures as described in Example 3. Sucrose was consumed almostcompletely by the end of the reaction. Aside from a viscous dextranproduct, 2918 gtf (SEQ ID NO:9) produced mostly fructose (˜50%), andsmall amounts of leucrose (˜5%) and glucose (˜1%). 2918 gtf (SEQ IDNO:9) produced a small amount of DP2-DP7 oligosaccharides (˜1%).

Thus, reactions comprising water, sucrose and an enzyme comprising SEQID NO:9 synthesized a gelling product, which is believed to be a dextranpolymer. Experimental results demonstrated that gtf 2920 likely hasglucosyltransferase activity.

Example 5 Expression of a Glucosyltransferase (2920) and Use Thereof toProduce a Gelling Dextran Product

This Example describes expression of a mature Streptococcus sobrinusglucosyltransferase (gtf) enzyme in B. subtilis. Also, this Exampleshows that this enzyme produces a gelling product, likely a dextran,when used is a reaction containing water and sucrose.

A glucosyltransferase gene, SsoGtf4, was identified from Streptococcussobrinus 813N. The nucleic acid sequence of this gene (positions 198 to4718 of GENBANK Accession No. AY966490) is set forth in SEQ ID NO:11 andencodes the protein sequence of SEQ ID NO:12 (GENBANK Accession No.AAX76986). At the N-terminus of the SsoGtf4 protein (SEQ ID NO:12) is asignal peptide of 41 amino acids, as predicted by the SIGNALP 4.0program. This indicates that SsoGtf4 (SEQ ID NO:12) is a secretedprotein. The mature, secreted form of the SsoGtf4 protein is hereinreferred to as 2920 gtf, and is set forth in SEQ ID NO:13.

The nucleotide sequence encoding 2920 gtf was optimized for expressionin B. subtilis. The optimized sequence (SEQ ID NO:14) was synthesized byGeneray (Shanghai, China), and inserted into plasmid p2JM103BBI,resulting in plasmid pZZHB584 (FIG. 2C). Plasmid pZZHB584 contains anaprE promoter operably linked to a sequence encoding (i) an aprE signalsequence used to direct heterologous protein (2920 gtf in this case)secretion in B. subtilis, (ii) Ala-Gly-Lys to facilitate the secretion,and (iii) 2920 gtf (SEQ ID NO:13) (i-iii are fused together in theamino-to-carboxy direction).

Plasmid pZZHB584 was transformed into B. subtilis cells for 2920 gtfexpression and purification (see General Methods).

The activity of 2920 gtf (SEQ ID NO:13) was determined in a 250-mLreaction at room temperature comprising 100 g/L sucrose, 20 mM sodiumphosphate buffer (pH 5.5), and 6.25 mL of enzyme stock. The reaction wascarried out at room temperature with shaking (150 rpm) for 6 days.

Samples (100 μL) were taken from the reaction at 0, 1, 3, 5, 24, 48, 72and 144 hour time points, respectively. Enzyme was deactivated byheating each sample at 80° C. for 10 minutes. Samples were diluted10-fold with water and centrifuged at 14000 rpm for 5 minutes.Supernatant (200 μL) was used for HPLC analysis.

The concentrations of sucrose, leucrose, glucose, fructose andoligosaccharides (DP2-DP7) in the gtf reaction were determined usingHPLC procedures as described in Example 3. Sucrose was consumed almostcompletely by the end of the reaction. Aside from a viscous dextranproduct, 2920 gtf (SEQ ID NO:13) produced mostly fructose (˜50%),leucrose (˜20%), and a small amount of glucose (˜3%). 2920 gtf (SEQ IDNO:13) produced a small amount of DP2-DP7 oligosaccharides (˜1%).

Thus, reactions comprising water, sucrose and an enzyme comprising SEQID NO:13 synthesized a gelling product, which is believed to be adextran polymer. Experimental results demonstrated that gtf 2920 likelyhas glucosyltransferase activity.

Example 6 Expression of a Glucosyltransferase (2921) and Use Thereof toProduce a Gelling Dextran Product

This Example describes expression of a mature Streptococcus downeiglucosyltransferase (gtf) enzyme in B. subtilis. Also, this Exampleshows that this enzyme produces a gelling product, likely a dextran,when used is a reaction containing water and sucrose.

A glucosyltransferase gene, SdoGtf7, was identified from Streptococcusdownei MFe28. The nucleic acid sequence of this gene (positions 16 to2375 of GENBANK Accession No. AB476746) is set forth in SEQ ID NO:15 andencodes the protein sequence of SEQ ID NO:16 (GENBANK Accession No.ZP_08549987.1). At the N-terminus of the SdoGtf7 protein (SEQ ID NO:16)is a signal peptide of 44 amino acids, as predicted by the SIGNALP 4.0program. This indicates that SdoGtf7 protein (SEQ ID NO:16) is asecreted protein. The mature, secreted form of the SdoGtf7 protein isherein referred to as 2921 gtf, and is set forth in SEQ ID NO:17.

The nucleotide sequence encoding 2921 gtf was optimized for expressionin B. subtilis. The optimized sequence (SEQ ID NO:18) was synthesized byGeneray (Shanghai, China), and inserted into plasmid p2JM103BBI,resulting in plasmid pZZHB585 (FIG. 2D). Plasmid pZZHB585 contains anaprE promoter operably linked to a sequence encoding (i) an aprE signalsequence used to direct heterologous protein (2921 gtf in this case)secretion in B. subtilis, (ii) Ala-Gly-Lys to facilitate the secretion,and (iii) 2921 gtf (SEQ ID NO:17) (i-iii are fused together in theamino-to-carboxy direction).

Plasmid pZZHB585 was transformed into B. subtilis cells for 2921 gtfexpression and purification (see General Methods).

The activity of 2921 gtf (SEQ ID NO:17) was determined in a 250-mLreaction at room temperature comprising 100 g/L sucrose, 20 mM sodiumphosphate buffer (pH 5.5), and 6.25 mL of enzyme stock. The reaction wascarried out at room temperature with shaking (150 rpm) for 8 days.

Samples (100 μL) were taken from the reaction at the reaction start andon 1, 2, 3, 6, 7 and 8 day time points, respectively. Enzyme wasdeactivated by heating each sample at 80° C. for 10 minutes. Sampleswere diluted 10-fold with water and centrifuged at 14000 rpm for 5minutes. Supernatant (200 μL) was used for HPLC analysis.

The concentrations of sucrose, leucrose, glucose, fructose andoligosaccharides (DP2-DP7) in the gtf reaction were determined usingHPLC procedures as described in Example 3. About 43% sucrose remained inthe reaction on day 8. Aside from a viscous dextran product, 2921 gtf(SEQ ID NO:17) produced mostly fructose (˜31%), leucrose (˜6%), andglucose (˜3%). No obvious production of DP2-DP7 oligosaccharides wasobserved.

Thus, reactions comprising water, sucrose and an enzyme comprising SEQID NO:17 synthesized a gelling product, which is believed to be adextran polymer. Experimental results demonstrated that gtf 2921 likelyhas glucosyltransferase activity.

Example 7 (Comparative) Production of Dextran Using CommerciallyAvailable Dextran Sucrase

This Example describes synthesizing dextran using a commerciallyavailable dextran sucrase in reactions comprising water and sucrose. Thedextran produced in this was analyzed in Example 8 in comparison to thegelling dextran products synthesized in Examples 1-6.

Reagents for preparing dextran sucrase reaction:

-   -   Sucrose (Sigma Prod. No. S-9378). 400 g/L stock solution was        prepared.    -   Sodium phosphate buffer stock (200 mM) (pH 5.5): prepare 250 mL        in water using sodium phosphate monobasic monohydrate (Sigma        Prod. No. S9638) and sodium phosphate dibasic heptahydrate        (Sigma Prod. No. S9390), accordingly.    -   Dextran sucrase, lyophilized powder, ≥100 units/mg protein, from        Leuconostoc mesenteroides (Sigma Prod. No. D9909).

A 50-mL reaction was prepared containing 20 mM sodium phosphate (pH5.5), 110 g/L sucrose, and 10 units of dextran sucrase fromSigma-Aldrich. The dextran sucrase was added last when preparing thereaction. The reaction was carried out in a 125-mL capped shake flask at26° C. with shaking (100 rpm) for 7 days. Samples (100 μL) of thereaction were taken at 0, 3, 6, 24, 48 and 168 hours, respectively. Thedextran sucrase was deactivated in each sample by heating at 80° C. for10 minutes. Each sample was then diluted 10-fold with water andcentrifuged at 14,000 rpm for 5 minutes, after which 200 μl ofsupernatant was used for HPLC analysis to measure sucrose consumptionduring the reaction.

The following HPLC conditions were applied for analyzing each sample:column (AMINEX HPX-87C carbohydrate column, 300×7.8 mm, Bio-Rad, No.125-0095), eluent (water), flow rate (0.6 mL/min), temperature (85° C.),refractive index detector. HPLC analysis of the samples indicatedsucrose consumption during the dextran sucrase reaction (FIG. 3). It isnotable that the sucrose consumption rate by the commercial dextransucrase was much slower compared to the sucrose consumption rate of gtf0768 (Example 2). Specifically, while gtf 0768 depleted most sucroseafter about 17-18 hours of reaction time (FIG. 1), commercial dextransucrase depleted only about 20% of sucrose within this same time period,and required about 168 hours to deplete all or most sucrose.

HPLC was also used to analyze other products of the reaction. Dextranyield was back-calculated by subtracting the amount of all othersaccharides left in the reaction from the amount of the startingsucrose. The back-calculated number was consistent with dextran dryweight analysis. Sucrose, leucrose, glucose, fructose, and DP2-7disaccharides were quantified by HPLC as described in Example 2. TheseHPLC analyses indicated that the saccharide products of the commercialdextran sucrase reaction consisted of 49% dextran, 0.3% sucrose, 44%fructose, 1% glucose, 5% leucrose, and 1% DP2-7 oligosaccharides.

The dextran produced in this Example was analyzed in Example 8 incomparison to the gelling dextran products synthesized in Examples 1-6.

Example 8 Viscosity of Dextran Samples

This Example describes measuring the viscosities of the dextran polymersproduced in Examples 1-7, as well as the viscosity of dextran obtainedfrom a commercial source. Viscosity measurements were made at variousshear rates.

Dextran polymer samples were prepared as described in Examples 1-7.Specifically, enzymatic reactions were conducted, after which polymerwas methanol-precipitated and washed with methanol (100%) four times,and then dried. Solutions (2 wt % and/or 3 wt %) of each sample wereprepared by adding the appropriate amount of polymer to de-ionized (DI)water. Each preparation was then mixed using a bench top vortexer untilpolymer was fully in solution. Each of these samples is referred to inTables 2 and 3 (below) as “After PPT” (after precipitation). A 2 wt %solution of dextran (Mw=956978) obtained from TCI America (Portland,Oreg.; catalogue No. D0061) was similarly prepared; this dextran isreferred to below as “commercial dextran”.

To determine the viscosity of each polymer solution at various shearrates, each solution was subjected to various shear rates using aviscometer while the temperature was held constant at 20° C. Also,polymer samples obtained directly, without precipitation, from each ofthe enzymatic reactions described in Examples 1-7 were subjected tovarious shear rates (referred to in Tables 2 and 3 as “Before PPT”). Theshear rate was increased using a gradient program which increased from0-10 rpm and the shear rate was increased by 0.17 (1/s) every 30seconds. The results of this experiment are listed in Table 2.

TABLE 2 Viscosity of Certain Dextran Solutions at Various Shear RatesViscosity Viscosity Viscosity Viscosity (cPs) @ (cPs) @ (cPs) @ (cPs) @Dextran Sample^(a) 0.17 rpm 1.03 rpm 2.62 rpm 4.22 rpm Gtf 0768 47976.1311376.70 12956.11 14390.76 (SEQ ID NO: 1) Before PPT (Example 2, 100 g/Lsucrose reaction) Gtf 0768 15778.40 6245.31^(b) 4119.58^(b) (SEQ IDNO: 1) After PPT - 3 wt % (Example 2, 100 g/L sucrose reaction) Gtf 07684091.84 3417.10 2874.10 (SEQ ID NO: 1) After PPT - 2 wt % (Example 2,100 g/L sucrose reaction) Gtf 2918 n/a^(b) n/a^(b) n/a^(b) (SEQ ID NO:9) Before PPT (Example 4) Gtf 2919 98864 38671 25580 (SEQ ID NO: 5)Before PPT (Example 3) Gtf 2920 3874.85 4205.66 4119.58^(b) (SEQ ID NO:13) Before PPT (Example 5) Gtf 2920 6168.76 3294.43 2288.24 (SEQ ID NO:13) After PPT - 3 wt % (Example 5) Gtf 2921 3533.86 2143.72 1748.95 (SEQID NO: 17) Before PPT (Example 6) Gtf 2921 4634.32 2780.4 1984.89 (SEQID NO: 17) After PPT - 3 wt % (Example 6) Commercial dextran 16759.42sucrase Before PPT (Example 7) ^(a)Polymer samples are listed accordingto the respective enzyme used to synthesize the sample. ^(b)Measurementwas outside the specification limits of the viscometer.

Polymer samples were also subjected to various higher shear rates usinga viscometer while the temperature was held constant at 20° C. The shearrate was increased using a gradient program which increased from 10-250rpm and the shear rate was increased by 7.36 (1/s) every 20 seconds. Theresults of this experiment are listed in Table 3.

TABLE 3 Viscosity of Certain Dextran Solutions at Various Shear RatesViscosity Viscosity Viscosity (cPs) @ (cPs) @ (cPs) @ Dextran Sample^(a)14.72 rpm 102.9 rpm 250 rpm Gtf 2918 (SEQ ID NO: 9) 149.95 69.68 48.97After PPT - 3 wt % (Example 4) Gtf 2919 (SEQ ID NO: 5) 80.82 41.23 29.49After PPT - 3 wt % (Example 3) 2 wt % Commercial 241.41 105.28 68.88dextran Commercial dextran sucrase 11.09^(b) 10.31^(b) 8.27 After PPT -2 wt % (Example 7) Viscosity Viscosity Viscosity (cPs) @ (cPs) @ (cPs) @14.11 rpm 98.69 rpm 162.1 rpm Gtf 0768 (SEQ ID NO: 1) 49.89 23.61 18.32After PPT - 2 wt % (Example 2, 400 g/L sucrose reaction) Gtf 0768 (SEQID NO: 1) 5.44 2.72 1.58 After PPT - 2 wt % (Example 2, 800 g/L sucrosereaction) ^(a)Polymer samples are listed according to the respectiveenzyme used to synthesize the sample. Alternatively, dextran obtainedfrom a commercial source was analyzed (“Commercial dextran”).^(b)Measurement was outside the specification limits of the viscometer.

These data demonstrate that solutions of the dextran product of aglucosyltransferase comprising SEQ ID NO:1 can in most cases exhibitincreased viscosity even after precipitation and resolvation, ascompared to the viscosities of commercially obtained dextran and thedextran product of a commercially obtained dextran sucrase. Thisobservation also appears to apply to the respective polymer products ofglucosyltransferases comprising SEQ ID NO:5, 9, 13, or 17.

It is also noteworthy that, based on Tables 2-3, as the amount ofsucrose in a gtf 0768 reaction is decreased from 800 g/L to 100 g/L, theviscosity of the dextran product appears to increase. Specifically,Table 3 indicates (at 14.11 rpm/2 wt % loading) viscosities of 5.44 cPsand 49.89 cPs for dextran products of reactions comprising 800 and 400g/L sucrose, respectively, and Table 2 (gtf 0768, 2 wt % loading) mayindicate a viscosity of about 957 cPs (exponential extrapolated at arotation of 14.11 rpm) for dextran product of a reaction comprising 100g/L sucrose. This result suggests that the viscosity of a dextranproduct can be controlled by modifying the level of sucrose initiallyprovided to reaction.

Example 9 Further Production and Analysis of Dextran Synthesized by Gtf0768

This Example is in addition to Example 2, describing another reactioncomprising water, sucrose and gtf 0768. Also, this Example providesadditional linkage and molecular weight analyses of the gelling productsynthesized by gtf 0768, showing that this product is a type of dextran.

Reagents for Preparing Gtf Reaction:

-   -   Sucrose (Sigma Prod. No. S-9378).    -   Sodium phosphate buffer stock (1 M, pH 6.5, Teknova Cat No:        S0276).    -   Gtf 0768 enzyme solution (cell lysate as prepared in Example 1).

Gtf Reaction Conditions:

A 50-mL reaction was prepared containing 20 mM sodium phosphate buffer(buffer was diluted 50-fold with ddH2O from 1 M stock, pH 6.5), 100 g/Lsucrose, and 0.1 mL of gtf 0768 enzyme solution. The reaction was shakenat 100 rpm in an incubator shaker (Innova, Model 4000) at 26° C. for 43hours; the reaction became viscous after about 24 hours.

The gtf enzyme was deactivated by heating the reaction at 80° C. for 10minutes. The deactivated viscous reaction was then mixed with 75 mL of100% methanol to precipitate the viscous product. A white precipitatewas formed. After carefully decanting the supernatant, the whiteprecipitate was washed twice with 75 mL of 100% methanol. The solidproduct was dried at 45° C. under vacuum in an oven for 48 hours.

Samples (1 mL) of the reaction were taken at 0, 0.5, 1, 2, and 24 hours,respectively. The gtf enzyme was deactivated in each sample by heatingat 80° C. for 10 minutes. Each sample was then diluted 10-fold withsterile water. 500 μL of diluted sample was transferred into acentrifuge tube filter (SPIN-X, 0.45-μm Nylon, 2.0 mL PolypropyleneTube, Costar #8170) and centrifuged at 12,000 rpm in a table centrifugefor 60 minutes, after which 200 μL of flowthrough was used for HPLCanalysis to measure sucrose consumption during the reaction. Thefollowing HPLC conditions were applied for analyzing each sample: column(AMINEX HPX-87C carbohydrate column, 300×7.8 mm, Bio-Rad, No. 125-0095),eluent (water), flow rate (0.6 mL/min), temperature (85° C.), refractiveindex detector. HPLC analysis of the samples indicated substantialsucrose consumption during the 0768 gtf reaction.

HPLC was also used to analyze other products of the reaction. Polymeryield was back-calculated by subtracting the amount of all othersaccharides left in the reaction from the amount of the startingsucrose. The back-calculated number was consistent with the viscousproduct dry weight analysis. Sucrose, leucrose, glucose and fructosewere quantified by HPLC with an HPX-87C column (HPLC conditions asdescribed above). DP2-7 oligosaccharides were quantified by HPLC withthe following conditions: column (AMINEX HPX-42A carbohydrate column,300×7.8 mm, Bio-Rad, No. 125-0097), eluent (water), flow rate (0.6mL/min), temperature (85° C.), refractive index detector. These HPLCanalyses indicated that the glucosyl-containing saccharide products ofthe 0768 gtf reaction consisted of 92.3% polymer product, 1.3% glucose,5.0% leucrose, and 1.4% DP2-7 oligosaccharides.

A sample of dry dextran powder product (˜0.2 g) of the above reactionwas used for molecular weight analysis. Molecular weight was determinedby a flow injection chromatographic method using an Alliance™ 2695separation module from Waters Corporation (Milford, Mass.) coupled withthree online detectors: a differential refractometer 2414 from Waters, aHeleos™-2 18-angle multiangle light scattering (MALS) photometer withquasielastic light scattering (QELS) detector from Wyatt Technologies(Santa Barbara, Calif.), and a ViscoStar™ differential capillaryviscometer from Wyatt. The dry dextran powder was dissolved at 0.5 mg/mLin aqueous Tris (Tris[hydroxymethyl]aminomethane) buffer (0.075 M)containing 200 ppm NaN₃. The dissolution of dextran was achieved byshaking overnight at 50° C. Two AQUAGEL-OH GUARD columns from AgilentTechnologies (Santa Clara, Calif.) were used to separate the dextranpolymer peak from the injection peak. The mobile base for this procedurewas the same as the dextran solvent, the flow rate was 0.2 mL/min, theinjection volume was 0.1 mL, and the column temperature was 30° C.Empower™ version 3 software from Waters was used for data acquisition,and Astra™ version 6 software from Wyatt was used for multidetector datareduction. It was determined from this work that the dextran polymerproduct had a weight-average molecular weight (Mw) of 1.022(+/−0.025)×10⁸ g/mol (i.e., roughly 100 million Daltons) (from MALSanalysis), a z-average radius of gyration of 243.33 (+/−0.42) nm (fromMALS analysis), and a z-average hydrodynamic radius of 215 nm (from QELSanalysis). It was also determined from QELS analysis that the dextranhas a standard deviation of particle size distribution (PSD) of about0.259, indicating that the dextran likely is polydisperse in terms ofhydrodynamic size.

For glycosidic linkage analysis purposes, a 50-mL gtf reaction wasprepared as described above in this Example, except that the reactiontime was 24 hours (reaction had become viscous). The gtf enzyme wasdeactivated by heating the reaction at 80° C. for 10 minutes. Thedeactivated viscous reaction was then placed into a regeneratedcellulose sturdy dialysis tubing with a molecular weight cut-off (MWCO)of 12-14 kDa (Spectra/Por® 4 Dialysis Tubing, Part No. 132706, SpectrumLaboratories, Inc.) and dialyzed against 4 L of filter water at roomtemperature over one week. Water was exchanged every day during thisdialysis. The dialyzed viscous reaction was then precipitated and driedas described above in this Example. About 0.2 g of dry powder wassubmitted for GC/MS linkage analysis.

Linkage analysis was performed according to methods described byPettolino et al. (Nature Protocols 7:1590-1607), which is incorporatedherein by reference. Briefly, a dry dextran sample was dissolved indimethyl sulfoxide (DMSO) or 5% lithium chloride in DMSO, then all freehydroxyl groups were methylated by sequential addition of a sodiumhydroxide/DMSO slurry followed by iodomethane. The methylated polymerwas then extracted into methylene chloride and hydrolyzed to monomericunits using aqueous trifluoroacetic acid (TFA) at 120° C. The TFA wasthen evaporated from the sample and reductive ring opening was doneusing sodium borodeuteride, which also labeled the reducing end with adeuterium atom. The hydroxyl groups created by hydrolyzing theglycosidic linkages were then acetylated by treating with acetylchloride and TFA at a temperature of 50° C. Finally, the derivatizingreagents were evaporated and the resulting methylated/acetylatedmonomers were reconstituted in acetonitrile and analyzed by gaschromatography with mass spectrometry (GC/MS) using a biscyanopropylcyanopropylphenyl polysiloxane column. The relative positioning of themethyl and acetyl functionalities, along with the deuterium label,yielded species that have distinctive retention time indices and massspectra that can be compared to published databases. In this way, thederivatives of the monomeric units indicated how each monomer wasoriginally linked in the dextran polymer and whether the monomer was abranch point. The results of analyzing these samples (dextran initiallydissolved in DMSO or DMSO/5% LiCl) are provided in Table 4.

TABLE 4 Linkage Profile of Gtf 0768 Dextran Product Wt %/Mol % ofGlucose Monomers in Dextran Sample 3-glc ^(a) 6-glc ^(b) 4-glc ^(c)3,6-glc ^(d) 2,6- + 4,6-glc ^(e) DMSO 0.4 90.2 0.4 8.3 0.7 DMSO/5% 0.989.3 0.4 8.0 1.4 LiCl ^(a) Glucose monomer linked at carbon positions 1and 3. ^(b) Glucose monomer linked at carbon positions 1 and 6. ^(c)Glucose monomer linked at carbon positions 1 and 4. ^(d) Glucose monomerlinked at carbon positions 1, 3 and 6. ^(e) Glucose monomer linked atcarbon positions 1, 2 and 6, or 1, 4 and 6.

In general, the results in Table 4 indicate that the dextran productanalyzed above comprises:

-   (i) about 87-93 wt % glucose linked only at positions 1 and 6;-   (ii) about 0.1-1.2 wt % glucose linked only at positions 1 and 3;-   (iii) about 0.1-0.7 wt % glucose linked only at positions 1 and 4;-   (iv) about 7.7-8.6 wt % glucose linked only at positions 1, 3 and 6;    and-   (v) about 0.4-1.7 wt % glucose linked only at (a) positions 1, 2 and    6, or (b) positions 1, 4 and 6.    Based on this information and some other studies (data not shown),    it is contemplated that this product is a branched structure in    which there are long chains (containing mostly or all    alpha-1,6-linkages) of about 20 DP in length (average) that    iteratively branch from each other (e.g., a long chain can be a    branch from another long chain, which in turn can itself be a branch    from another long chain, and so on). The branched structure also    appears to comprise short branches from the long chains; these short    chains are believed to be 1-3 DP in length and mostly comprise    alpha-1,3 and -1,4 linkages, for example. Branch points in the    dextran, whether from a long chain branching from another long    chain, or a short chain branching from a long chain, appear to    comprise alpha-1,3, -1,4, or -1,2 linkages off of a glucose involved    in alpha-1,6 linkage. Roughly 25% of all the branch points of the    dextran branched into a long chain.

Thus, reactions comprising water, sucrose and an enzyme comprising SEQID NO:1 synthesized a very large gelling dextran product, as determinedby the product's high Mw and predominant alpha-1,6 glucosidic linkageprofile.

Example 10 Formulation Comprising Dextran Synthesized by Gtf 0768

This Example discloses a formulation comprising the dextran product ofgtf 0768. This formulation was shown to have better sensorycharacteristics (or “feel”) compared to formulations comprising certaincompounds (xanthan gum, Carbopol®) commonly used for providing viscosityto certain consumer products (e.g., personal care compositions such aslotion).

Three different emulsions were prepared and compared against each otherin a skinfeel study, as follows.

Dextran-Based Emulsion:

Dextran was produced using gtf 0768 (comprising SEQ ID NO:1) in areaction similar to the reaction disclosed in Example 9. At roomtemperature, polysorbate 80, sorbitan monooleate and mineral oil (PhaseB, Table 5) were combined in a small vessel, and mixed by hand untilhomogeneous. Phase B was slowly added to water (Phase A, Table 5) undermoderate propeller mixing. The mixture was homogenized at 5000-9000 rpmfor approximately 5-10 minutes. Dextran (Phase C, Table 5) was thenadded under moderate propeller mixing. Germaben® II (Phase D, Table 5)was then added as a preservative under moderate propeller mixing. Thedextran could optionally have been pre-hydrated using a portion of thewater from phase A.

TABLE 5 Dextran-Based Emulsion % wt % wt % Ingredients Activity(Desired) (Neat) Grams Phase A Water (deionized) 73.50 73.50 Phase BPolysorbate 80 100.00 2.43 2.43 2.43 Sorbitan Monooleate 100.00 2.572.57 2.57 Mineral Oil 100.00 20.00 20.00 20.00 Phase C Dextran 100.001.00 1.00 1.00 Phase D Germaben ® II 100.00 0.50 0.50 0.50 100.00 100.00

Xanthan Gum-Based Emulsion (Control 1):

At room temperature, xanthan gum and water (Phase A, Table 6) werecombined under moderate propeller mixing until homogeneous. Polysorbate80, sorbitan monooleate and mineral oil (Phase B, Table 6) were combinedin a small vessel, and mixed by hand until homogeneous. Phase B wasslowly added to Phase A under moderate propeller mixing. The mixture washomogenized at 5000-9000 rpm for approximately 5-10 minutes. Germaben®II (Phase C, Table 6) was then added as a preservative under moderatepropeller mixing.

TABLE 6 Xanthan Gum-Based Emulsion % wt % wt % Ingredients Activity(Desired) (Neat) Grams Phase A Water (deionized) 74.00 74.00 Xanthan Gum100.00 0.50 0.50 0.50 Phase B Polysorbate 80 100.00 2.43 2.43 2.43Sorbitan Monooleate 100.00 2.57 2.57 2.57 Mineral Oil 100.00 20.00 20.0020.00 Phase C Germaben ® II 100.00 0.50 0.50 0.50 100.00 100.00

Carbopol® Ultrez 10-Based Emulsion (Control 2):

At room temperature, Carbopol® Ultrez 10 and water (Phase A, Table 7)were combined under moderate propeller mixing until homogeneous.Polysorbate 80, sorbitan monooleate and mineral oil (Phase B, Table 7)were combined in a small vessel, and mixed by hand until homogeneous.Phase B was slowly added to Phase A under moderate propeller mixing. Themixture was homogenized at 5000-9000 rpm for approximately 5-10 minutes.Germaben® II (Phase C, Table 7) was then added as a preservative undermoderate propeller mixing. A 20-wt % solution of sodium hydroxide wasused to neutralize the emulsion to pH 5.5.

TABLE 7 Carbopol ® Ultrez 10-Based Emulsion % wt % wt % IngredientsActivity (Desired) (Neat) Grams Phase A Water (deionized) 74.00 74.00Carbopol ® Ultrez 10 100.00 0.50 0.50 0.50 Phase B Polysorbate 80 100.002.43 2.43 2.43 Sorbitan Monooleate 100.00 2.57 2.57 2.57 Mineral Oil100.00 20.00 20.00 20.00 Phase C Germaben ® II 100.00 0.50 0.50 0.50100.00 100.00

Skinfeel Analysis and Results:

A double-blind, skinfeel analysis was performed according to ASTME1490-3 (“Standard Practice for Descriptive Skinfeel Analysis of Creamsand Lotions”, ASTM International, West Conshohocken, Pa., 2003, DOI:10.15201E1490-03, incorporated herein by reference) to compare each ofthe above emulsions. The primary attributes evaluated in this study wererub-out sliminess, afterfeel stickiness, pick-up stringiness and pick-upstickiness. Panelists assessed attributes on a scale from 1-5, where 1exhibits the least of the attribute and 5 exhibits the most of theattribute. The results are reported in Table 8 below as an average valueof the panelists' ratings for each attribute. The sum average of thesevalues (Σ, Table 8) indicates that the overall sensory experience foremulsions (e.g., lotions) produced with dextran as presently disclosedexceeds the results of similar emulsions produced with either xanthangum or Carbopol® Ultrez 10.

TABLE 8 Carbopol ® Ultrez 10-Based Emulsion Average Rating XanthanCarbopol ® Skinfeel Attribute Dextran Gum Ultrez 10 Rub-Out Sliminess 23 2 Afterfeel Stickiness 2 2 3 Pick-Up Stringiness 1 3 3 Pick-UpStickiness 2 3 2 Σ 7 11 10

It is noteworthy that the dextran-containing emulsion scored better thanthe control emulsions in the skinfeel analysis, especially since therewas two-times the amount of dextran (1 wt %) in the emulsion compared tothe amount of xanthan gum (0.5 wt %) or Carbopol® Ultrez 10 (0.5 wt %)in the control emulsions.

Thus, dextran produced by gtf 0768 (comprising SEQ ID NO:1) can besuitable for use in compositions where enhanced sensory characteristicsare desirable, such as in personal care and food products, for example.

Example 11 Dextran-Comprising Cleanser with Suspended Particles

This Example discloses a cleanser comprising the dextran product of gtf0768. Jojoba ester beads could be suspended in this composition,indicating that the dextran can function as a dispersant.

Dextran was produced using gtf 0768 (comprising SEQ ID NO:1) in areaction similar to the reaction disclosed in Example 9. At roomtemperature water, dextran, glycerin, polysorbate 20, cocamidopropylbetaine, PPG-2 hydroxyethyl cocamide and disodium EDTA were combinedaccording to the formulation in Table 9, and mixed by hand untilhomogeneous. Jojoba beads were then added and mixing was continued untilthe beads were homogeneously dispersed. The dextran could optionallyhave been pre-hydrated using a portion of the water component.

TABLE 9 Dextran-Based Jojoba Bead Suspension % wt % wt % IngredientActivity (Desired) (Neat) Grams Water (deionized) 22.95 22.95 Dextran100 5 5 5 Glycerin 100 10 10 10 Polysorbate 20 100 5.25 5.25 5.25Cocamidopropyl Betaine 35.97 20 55.6 55.6 PPG-2 Hydroxyethyl 100 1 1 1Cocamide Disodium EDTA 100 0.1 0.1 0.1 Jojoba Ester Beads 100 0.1 0.10.1 100 100

Thus, dextran produced by gtf 0768 (comprising SEQ ID NO:1) can be usedas a dispersant in aqueous compositions such as certain personal careproducts.

What is claimed is:
 1. A composition comprising an isolated dextran,wherein said dextran comprises: (i) about 87-93 wt % glucose linked atpositions 1 and 6; (ii) about 0.1-1.2 wt % glucose linked at positions 1and 3; (iii) about 0.1-0.7 wt % glucose linked at positions 1 and 4;(iv) about 7.7-8.6 wt % glucose linked at positions 1, 3 and 6; and (v)about 0.4-1.7 wt % glucose linked at: (a) positions 1, 2 and 6, or (b)positions 1, 4 and 6; wherein the weight-average molecular weight (Mw)of said dextran is about 50-200 million Daltons, the z-average radius ofgyration of said dextran is about 200-280 nm, and the dextran is aproduct of an isolated glucosyltransferase enzyme comprising an aminoacid sequence that is at least 90% identical to SEQ ID NO:1 or SEQ IDNO:2.
 2. The composition of claim 1, wherein the dextran comprises: (i)about 89.5-90.5 wt % glucose linked at positions 1 and 6; (ii) about0.4-0.9 wt % glucose linked at positions 1 and 3; (iii) about 0.3-0.5 wt% glucose linked at positions 1 and 4; (iv) about 8.0-8.3 wt % glucoselinked at positions 1, 3 and 6; and (v) about 0.7-1.4 wt % glucoselinked at: (a) positions 1, 2 and 6, or (b) positions 1, 4 and
 6. 3. Thecomposition of claim 1, wherein the dextran comprises chains iterativelylinked together within a branching structure, wherein said chains aresimilar in length and comprise substantially alpha-1,6-glucosidiclinkages.
 4. The composition of claim 3, wherein the average length ofthe chains is about 10-50 monomeric units.
 5. The composition of claim1, wherein the dextran is a product of an isolated glucosyltransferaseenzyme comprising an amino acid sequence that is at least 95% identicalto SEQ ID NO:1 or SEQ ID NO:2.
 6. The composition of claim 1, whereinthe composition is an aqueous composition having a viscosity of at leastabout 25 centipoise.
 7. The composition of claim 1, wherein the Mw ofthe dextran is about 80-120 million Daltons.
 8. The composition of claim1, wherein the z-average radius of gyration of said dextran is about230-250 nm.
 9. The composition of claim 1, wherein the composition is inthe form of a food product, personal care product, pharmaceuticalproduct, household product, or industrial product.
 10. The compositionof claim 9, wherein the composition is in the form of a confectionery.11. A method for increasing the viscosity of an aqueous composition, themethod comprising: contacting dextran according to claim 1 with theaqueous composition, wherein the viscosity of the aqueous composition isincreased by said dextran compared to the viscosity of the aqueouscomposition before the contacting step.
 12. A method of treating amaterial, said method comprising: contacting the material with anaqueous composition comprising dextran according to claim 1.